Instant beverage powder based on blg

ABSTRACT

The present invention relates to an instant beverage powder product and a method for preparing the instant beverage powder product, a liquid food product produced from the instant beverage powder and a method for preparing the liquid food, use of the liquid food, and a kit comprising the instant beverage powder product.

FIELD OF THE INVENTION

The present invention relates to an instant beverage powder product anda method for preparing the instant beverage powder product, a liquidfood product produced from the instant beverage powder and a method forpreparing the liquid food, use of the liquid food, and a kit comprisingthe instant beverage powder product.

BACKGROUND

Nutritional supplements comprising milk serum proteins are commonly usedfor muscle synthesis, for weight control and for maintaining muscle andbody weight. Nutritional supplements are targeted different kinds ofconsumers, e.g. sportsmen/women, athletes, children, elderly people andpatients with or at risk of malnutrition, and/or with increased proteinneeds. Thus, the consumer perception of the nutritional supplement is ofgreat importance, as the consumer should feel for drinking the product.

Milk serum proteins can be isolated from milk serum or whey. Wheytypically comprises a mixture of beta-lactoglobulin (BLG),alpha-lactalbumin (ALA), serum albumin and immunoglobulins, of which BLGis the most dominant. Whey protein concentrates (WPC) thus comprise amixture of these proteins. Whey protein isolates (WPI) contain less fatand lactose than WPC. Isolation of beta-lactoglobulin (BLG) from milkserum or whey is the subject of a number of publications and typicallyinvolves multiple separation steps and often chromatographic techniquesto arrive at a purified beta-lactoglobulin product.

International patent application WO2002/056707 (Nestle) concerns abalanced powder blend composition with at least one fat or oil source,at least one carbohydrate source, and at least on protein source, isdescribed. This composition is advantageously added to a food tosupplement the nutritional value of the food, but without substantiallyaltering the taste of the food.

WO 2018/115520 A1 discloses a method of producing edible isolatedbeta-lactoglobulin compositions and/or compositions containingcrystallised beta-lactoglobulin based on crystallisation of BLG insalting-in mode. The crystallised BLG may subsequently be separated fromthe remaining mother liquor.

WO 2011/112695 A1 discloses nutritional compositions and methods ofmaking and using the nutritional compositions. The nutritionalcompositions comprise whey protein micelles and leucine and provide asufficient amount of leucine to improve protein synthesis in humans,while also maintaining a low-viscosity fluid matrix and acceptableorganoleptic properties.

WO2011/051436 A1 discloses an at least partially transparent compositionintended for human or animal consumption and relates to the packaging ofsuch compositions. One embodiment of the present invention relates to anat least partially transparent container containing an at leastpartially transparent aqueous non-alcoholic composition. The containercomprises at least one polarizer that makes liquid crystals present inthe composition visible.

WO2004/049819 A2 discloses a method for improving the functionalproperties of globular proteins, comprising the steps of providing asolution of one or more globular proteins, in which solution theprotein(s) is/are at least partially aggregated in fibrils; andperforming one or more of the following steps in random order:increasing the pH; increasing the salt concentration; concentrating thesolution; and changing the solvent quality of the solution. Preferably,the solution of the one or more globular protein is provided by heatingat a low pH or the addition of a denaturing agent. Disclosed is also theprotein additive thus obtained, the use thereof for food and non-foodapplications and to the food and non-food products containing theprotein additive.

WO 2010/037736 A1 discloses isolation of whey proteins and thepreparation of a whey product and a whey isolate. In particular thepresent invention relates to the isolation of a β-lactoglobulin productand the isolation of an α-enriched whey protein isolate from wheyobtained from an animal. The α-enriched whey protein isolate provided bythe present invention is besides from being low in β-lactoglobulin alsohigh in α-lactalbumin and immunoglobulin G.

FR 2 296 428 discloses protein compositions for dietetic and therapeuticuse based on lactoserum proteins obtained by any known separationprocess. The compositions can be used for the treatment or prophylaxisof digestive disorders in infants and adults (e.g. diarrhoea), toincrease resistance to intestinal infections, and to treat certainmetabolic disorders (e.g. hyper-phylalaninaemia). They can also be useddermatologically or cosmetically, and can form part of a low-proteindiet.

SUMMARY OF THE INVENTION

The inventors have provided instant beverage powder products with a highcontent of BLG. The products are shelf stable, while at the same timeresulting in food products that are appetizing; i.e. the appearance andtaste of the product is appealing to the customer.

Thus, an aspect of the invention pertains to an instant beverage powdercomprising at least 1% w/w BLG, preferably at least 5%, wherein:

-   -   i. the crystallinity of BLG is at least 20%, preferably at least        40%, and/or    -   ii. at least 85% w/w of the total amount of protein is comprised        by BLG,        and furthermore comprising at least one additional ingredient        selected from the group consisting of vitamins, flavouring        agent, colouring agent, minerals, sweeteners, antioxidants, food        acid, lipids, carbohydrate, prebiotics, probiotics, anti-foaming        agents and non-whey protein.

Another aspect of the invention pertains to a method for preparing aninstant beverage powder comprising BLG and at least one optionalingredient, said method comprising blending a dry BLG isolate with theleast one additional ingredient selected from the group consisting ofvitamins, flavouring agent, colouring agent, minerals, sweeteners,antioxidants, food acid, lipids, carbohydrate, prebiotics, probiotics,antifoaming agents and non-whey protein to obtain an instant beveragepowder.

Yet an aspect of the invention pertains to a liquid food productcomprising a liquid and the powder according to the invention.

A further aspect of the invention pertains to a method for preparing aliquid food product according to the invention, said method comprising

-   -   i. Adding an instant beverage powder according to the invention,    -   ii. Optionally adding at least one further ingredient, and    -   iii. Mixing the powder and liquid obtained to form a uniform        mixture.

A further aspect of the invention pertains to an instant beverage powderaccording to the invention, for use as a nutritional supplement.

A further aspect of the invention pertains to a kit comprising thepowder according to the invention,

-   -   i. a tool for measuring said powder, and    -   ii. a container having a lid for opening and closing the        container,    -   wherein said container is for mixing said powder with a liquid        to form a food product, and said container is adapted for        drinking the food product directly from the container.

BRIEF DESCRIPTION OF THE FIGURES

FIG. 1 is a microscope photo of the BLG crystals recovered from feed 3of Example 3 of the PCT application PCT/EP2017/084553.

FIG. 2 shows a microscope photo of the BLG crystals, both whole andfragmented, obtained from feed 2 of Example 3 of the PCT applicationPCT/EP2017/084553.

FIG. 3 is a photo of test tubes containing sub-samples of the six lowphosphorous beverages as prepared in example 5.

DEFINITIONS

In the context of the present invention, the term “beta-lactoglobulin”or “BLG” pertains to beta-lactoglobulin from mammal species, e.g. innative, unfolded and/or glycosylated forms and includes the naturallyoccurring genetic variants. The term furthermore includes aggregatedBLG, precipitated BLG and crystalline BLG. When referring to the amountof BLG reference is made to the total amount of BLG including aggregatedBLG. The total amount of BLG is determined according to Example 1.31.The term “aggregated BLG” pertains to BLG which is at least partiallyunfolded and which furthermore has aggregated with other denatured BLGmolecules and/or other denatured whey proteins, typically by means ofhydrophobic interactions and/or covalent bonds.

BLG is the most predominant protein in bovine whey and milk serum andexists in several genetic variants, the main ones in cow milk beinglabelled A and B. BLG is a lipocalin protein, and can bind manyhydrophobic molecules, suggesting a role in their transport. BLG hasalso been shown to be able to bind iron via siderophores and might havea role in combating pathogens. A homologue of BLG is lacking in humanbreast milk.

Bovine BLG is a relatively small protein of approx. 162 amino acidresidues with a molecular weight of approx. 18.3-18.4 kDa. Underphysiological conditions, it is predominantly dimeric, but dissociatesto a monomer below about pH 3, preserving its native state as determinedusing Nuclear Magnetic Resonance spectroscopy. Conversely, BLG alsooccurs in tetrameric, octameric and other multimeric aggregation formsunder a variety of natural conditions.

In the context of the present invention, the term “non-aggregatedbeta-lactoglobulin” or “non-aggregated BLG” also pertains tobeta-lactoglobulin from mammal species, e.g. in native, unfolded and/orglycosylated forms and includes the naturally occurring geneticvariants. However, the term does not include aggregated BLG,precipitated BLG or crystallised BLG. The amount or concentration ofnon-aggregated BLG is determined according to Example 1.6.

The percentage of non-aggregated BLG relative to total BLG is determinedby calculate (m_(total BLG)−m_(non-aggregate BLG))/m_(total BLG)*100%.m_(total BLG) is the concentration or amount of BLG determined accordingto Example 1.31 and m_(non-aggregated BLG) is the concentration oramount of non-aggregated BLG determined according to Example 1.6.

In the context of the present invention, the term “crystal” pertains toa solid material whose constituents (such as atoms, molecules or ions)are arranged in a highly ordered microscopic structure, forming acrystal lattice that extends in all directions.

In the context of the present invention, the term “BLG crystal” pertainsto protein crystals that primarily contain non-aggregated and preferablynative BLG arranged in a highly ordered microscopic structure, forming acrystal lattice that extends in all directions. The BLG crystals maye.g. be monolithic or polycrystalline and may e.g. be intact crystals,fragments of crystals, or a combination thereof. Fragments of crystalare e.g. formed when intact crystals are subjected to mechanical shearduring processing. Fragments of crystals also have the highly orderedmicroscopic structure of crystal but may lack the even surface and/oreven edges or corners of an intact crystal. See e.g. FIG. 1 for anexample of many intact BLG crystals and FIG. 2 for an example offragments of BLG crystals. In both cases, the BLG crystal or crystalfragments can be identified visually as well-defined, compact andcoherent structures using light microscopy. BLG crystal or crystalfragments are often at least partially transparent. Protein crystals arefurthermore known to be birefringent and this optical property can beused to identify unknown particles having a crystal structure.Non-crystalline BLG aggregates, on the other hand, often appear aspoorly defined, non-transparent, and as open or porous lumps ofirregular size.

In the context of the present invention, the term “crystallise” pertainsto the formation of protein crystals. Crystallisation may e.g. happenspontaneously or be initiated by the addition of crystallisation seeds.

In the context of the present invention, the term “edible composition”pertains to a composition that is safe for human consumption and use asa food ingredient and that does not contain problematic amounts of toxiccomponents, such as toluene or other unwanted organic solvents.

In the context of the present invention, the term “ALA” or“alpha-lactalbumin” pertains to alpha-lactalbumin from mammal species,e.g. in native and/or glycosylated forms and includes the naturallyoccurring genetic variants. The term furthermore includes aggregated ALAand precipitated BLG. When referring to the amount of ALA reference ismade to the total amount of ALA including e.g. aggregated ALA. The totalamount of ALA is determined according to Example 1.31. The term“aggregated ALA” pertains to ALA which typically is at least partiallyunfolded and which furthermore has aggregated with other denatured ALAmolecules and/or other denatured whey proteins, typically by means ofhydrophobic interactions and/or covalent bonds.

Alpha-lactalbumin (ALA) is a protein present in the milk of almost allmammalian species. ALA forms the regulatory subunit of the lactosesynthase (LS) heterodimer and β-1,4-galactosyltransferase (beta4Gal-T1)forms the catalytic component. Together, these proteins enable LS toproduce lactose by transferring galactose moieties to glucose. One ofthe main structural differences with beta-lactoglobulin is that ALA doesnot have any free thiol group that can serve as the starting-point for acovalent aggregation reaction.

In the context of the present invention, the term “non-aggregated ALA”also pertains to ALA from mammal species, e.g. in native, unfoldedand/or glycosylated forms and includes the naturally occurring geneticvariants. However, the term does not include aggregated ALA orprecipitated ALA. The amount or concentration of non-aggregated BLG isdetermined according to Example 1.6.

The percentage of non-aggregated ALA relative to total ALA is determinedby calculate (m_(total ALA)−m_(non-aggregate ALA))/m_(total ALA)*100%.m_(total ALA) is the concentration or amount of ALA determined accordingto Example 1.31 and m_(non-aggregated ALA) is the concentration oramount of non-aggregated ALA determined according to Example 1.6.

In the context of the present invention, the term “caseinomacropeptide”or “CMP” pertains to the hydrophilic peptide, residue 106-169,originated from the hydrolysis of “κ-CN” or “kappa-casein” from mammalspecies, e.g. in native and/or glycosylated forms and includes thenaturally occurring genetic variants, by an aspartic proteinase, e.g.chymosin.

In the context of the present invention, the term “BLG isolate” means acomposition that contains BLG in an amount of at least 85% w/w relativeto total protein. A BLG isolate preferably has a total protein contentof a least 30% w/w, and preferably at least 80% w/w relative to totalsolids.

In the context of the present invention, the term “BLG isolate powder”pertains to a BLG isolate in powder form and preferably a free-flowingpowder.

In the context of the present invention, the term “BLG isolate liquid”pertains to a BLG isolate in liquid form and preferably an aqueousliquid.

The term “whey” pertains to the liquid phase that is left after thecasein of milk has been precipitated and removed. Casein precipitationmay e.g. be accomplished by acidification of milk and/or by use ofrennet enzyme. Several types of whey exist, such as “sweet whey”, whichis the whey product produced by rennet-based precipitation of casein,and “acid whey” or “sour whey”, which is the whey product produced byacid-based precipitation of casein. Acid-based precipitation of caseinmay e.g. be accomplished by addition of food acids or by means ofbacterial cultures.

The term “milk serum” pertains to the liquid which remains when caseinand milk fat globules have been removed from milk, e.g. bymicrofiltration or large pore ultrafiltration. Milk serum may also bereferred to as “ideal whey”.

The term “milk serum protein” or “serum protein” pertains to the proteinwhich is present in the milk serum.

In the context of the present invention, the term “whey protein”pertains to protein that is found in whey or in milk serum. Whey proteinmay be a subset of the protein species found in whey or milk serum, andeven a single whey protein species or it may be the complete set ofprotein species found in whey or/and in milk serum.

In the context of the present invention, the main non-BLG proteins of astandard whey protein concentrate from sweet whey are ALA, CMP, bovineserum albumin, immunoglobulin, osteopontin, lactoferrin, andlactoperoxidase. In the context of the present invention, the weightpercentages of the main non-BLG whey proteins of a standard whey proteinconcentrate from sweet whey are:

ALA in an amount of 18% w/w relative to total protein,CMP in an amount of 18% w/w relative to total protein,BSA in an amount of 4% w/w relative to total protein,Casein species in an amount of 5% w/w relative to total protein,Immunoglobulin in an amount of 6% w/w relative to total protein,Osteopontin in an amount of 0.5% w/w relative to total protein,Lactoferrin in an amount of 0.1% w/w relative to total protein, andLactoperoxidase in an amount of 0.1% w/w relative to total protein.

The term casein pertains to casein protein found in milk and encompassesboth native micellar casein as found in raw milk, the individual caseinspecies, and caseinates.

In the context of the present invention, a liquid which is“supersaturated” or “supersaturated with respect to BLG” contains aconcentration of dissolved, non-aggregated BLG which is above thesaturation point of non-aggregated BLG in that liquid at the givenphysical and chemical conditions. The term “supersaturated” iswell-known in the field of crystallisation (see e.g. Gérand Coquerela,“Crystallization of molecular systems from solution: phase diagrams,supersaturation and other basic concepts”, Chemical Society Reviews, p.2286-2300, Issue 7, 2014) and supersaturation can be determined by anumber of different measurement techniques (e.g. by spectroscopy orparticle size analysis). In the context of the present invention,supersaturation with respect to BLG is determined by the followingprocedure.

Procedure for Testing Whether a Liquid at a Specific Set of Conditionsis Supersaturated with Respect to BLG:a) Transfer a 50 ml sample of the liquid to be tested to a centrifugetube (VWR Catalogue no. 525-0402) having a height of 115 mm, an insidediameter of 25 mm and a capacity of 50 mL. Care should be taken to keepthe sample and subsequent fractions thereof at the original physical andchemical conditions of the liquid during steps a)-h).b) The sample is immediately centrifuged at 3000 g for 3.0 minutes withmax. 30 seconds acceleration and max 30 seconds deceleration.c) Immediately after the centrifugation, transfer as much as possible ofthe supernatant (without disturbing the pellet if a pellet has formed)to a second centrifuge tube (same type as in step a)d) Take a 0.05 mL subsample of the supernatant (subsample A)e) Add 10 mg of BLG crystals (at least 98% pure, non-aggregated BLGrelative to total solids) having a particle size of at most 200 micronto a second centrifuge tube and agitate the mixture.f) Allow the second centrifuge tube to stand for 60 minutes at theoriginal temperature.g) Immediately after step f), centrifuge the second centrifuge tube at500 g for 10 minutes and then take another 0.05 mL subsample of thesupernatant (subsample B).h) Recover the centrifugation pellet of step g) if there is one,resuspend it in milliQ water and immediately inspect the suspension forpresence of crystals that are visible by microscopy.i) Determine the concentration of non-aggregated BLG in subsamples A andB using the method outlined in Example 1.6—the results are expressed as% BLG w/w relative to the total weight of the subsamples. Theconcentration of non-aggregated BLG of subsample A is referred to asC_(BLG,A), and the concentration of non-aggregated BLG of subsample B isreferred to as C_(BLG,B).j) The liquid from which the sample of step a) was taken wassupersaturated (at the specific conditions) if C_(BLG,B) is lower thanC_(BLG,A) and if crystals are observed in step i).

In the context of the present invention, the terms “liquid” and“solution” encompass both compositions that are free of particulatematter and compositions that contain a combination of liquid and solidand/or semi-solid particles, such as e.g. protein crystals or otherprotein particles. A “liquid” or a “solution” may therefore be asuspension or even a slurry. However, a “liquid” and “solution” arepreferably pumpable.

In the context of the present invention, the terms “whey proteinconcentrate” (WPC) and “serum protein concentrate” (SPC) pertain to dryor aqueous compositions which contain a total amount of protein of20-89% w/w relative to total solids.

A WPC or an SPC preferably contains:

20-89% w/w protein relative to total solids,15-70% w/w BLG relative to total protein,8-50% w/w ALA relative to total protein, and0-40% w/w CMP relative to protein.

Alternatively, but also preferred, a WPC or an SPC may contain:

20-89% w/w protein relative to total solids,15-90% w/w BLG relative to total protein,4-50% w/w ALA relative to total protein, and0-40% w/w CMP relative to protein.

Preferably, a WPC or an SPC contains:

20-89% w/w protein relative to total solids,15-80% w/w BLG relative to total protein,4-50% w/w ALA relative to total protein, and0-40% w/w CMP relative to protein.

More preferably a WPC or an SPC contains:

70-89% w/w protein relative to total solids,30-90% w/w BLG relative to total protein,4-35% w/w ALA relative to total protein, and0-25% w/w CMP relative to protein.

SPC typically contain no CMP or only traces of CMP.

The terms “whey protein isolate” (WPI) and “serum protein isolate” (SPI)pertain to dry or aqueous compositions which contain a total amount ofprotein of 90-100% w/w relative to total solids.

A WPI or an SPI preferably contains:

90-100% w/w protein relative to total solids,15-70% w/w BLG relative to total protein,8-50% w/w ALA relative to total protein, and0-40% w/w CMP relative to total protein.

Alternatively, but also preferred, a WPI or an SPI may contain:

90-100% w/w protein relative to total solids,30-95% w/w BLG relative to total protein,4-35% w/w ALA relative to total protein, and0-25% w/w CMP relative to total protein.

More preferably a WPI or an SPI may contain:

90-100% w/w protein relative to total solids,30-90% w/w BLG relative to total protein,4-35% w/w ALA relative to total protein, and0-25% w/w CMP relative to total protein.

SPI typically contain no CMP or only traces of CMP.

In the context of the present invention, the term “additional protein”means a protein that is not BLG. The additional protein that is presentin the whey protein solution typically comprises one or more of thenon-BLG proteins that are found in milk serum or whey. Non-limitingexamples of such proteins are alpha-lactalbumin, bovine serum albumin,immunoglobulines, caseinomacropeptide (CMP), osteopontin, lactoferrin,and milk fat globule membrane proteins.

The terms “consists essentially of” and “consisting essentially of” meanthat the claim or feature in question encompasses the specifiedmaterials or steps and those that do not materially affect the basic andnovel characteristic(s) of the claimed invention.

In the context of the present invention, the phrase “Y and/or X” means“Y” or “X” or “Y and X”. Along the same line of logic, the phrase “n₁,n₂, . . . , n_(i−1), and/or n_(i)” means “n₁” or “n₂” or . . . or“n_(i−1)” or “n_(i)” or any combination of the components: n₁, n₂, . . .n_(i−1), and n_(i).

In the context of the present invention, the term “dry” or “dried” meansthat the composition or product in question comprises at most 10% w/wwater, preferably at most 6% w/w and more preferably even less.

In the context of the present invention, the term “physical microbialreduction” pertains to physical interaction with a composition whichresults in reduction of the total amount of viable microorganisms of thecomposition. The term does not encompass addition of chemicals thatresult in killing of microorganisms. The term furthermore does notencompass the heat exposure to which the atomized droplets of liquid areexposed to during spray-drying but include possible pre-heating prior tospray-drying.

In the context of the present invention, the pH of a powder refers tothe pH of 10 g of the powder mixed into 90 g demineralised water and ismeasured according to Example 1.16.

In the context of the present invention, the weight percentage (% w/w)of a component of a certain composition, product, or material means theweight percentage of that component relative to the weight of thespecific composition, product, or material unless another reference (e.gtotal solids or total protein) is specifically mentioned.

In the context of the present invention, the process step“concentration” and the verb “concentrate” pertain to concentration ofprotein and encompass both concentration of protein on total solidsbasis and concentration of protein on a total weight basis. This meanse.g. that concentration does not necessarily require that the absoluteconcentration w/w of protein of a composition increases as long at thecontent of protein increases relative to total solids.

In the context of the present invention, the term “weight ratio” betweencomponent X and component Y means the value obtained by the calculationm_(X)/m_(Y) wherein m_(X) is the amount (weight) of components X andm_(Y) is the amount (weight) of components Y.

In the context of the present invention, the term “at leastpasteurisation” pertains to a heat-treatment which has microbial killingeffect equal to or higher than a heat-treatment of 70 degrees C. for 10seconds. The reference for determining the bacteria killing effect is E.coli O157:H7.

In the context of the present invention, the term “whey protein feed”pertains to whey protein source from which the liquid BLG isolate isderived. The whey protein feed has a lower content of BLG relative tototal protein than the liquid BLG isolate and is typically a WPC, a WPI,an SPC or an SPI.

In the context of the present invention, the term “BLG-enrichedcomposition” pertains to the BLG-enriched composition resulting fromisolating BLG from the whey protein feed. The BLG-enriched compositiontypically comprises the same whey proteins as the whey protein feed butBLG is present in significantly higher concentration relative to totalprotein than in whey protein feed. The BLG-enriched composition may e.g.be prepared from the whey protein feed by chromatography, proteincrystallisation and/or membrane-based protein fractionation. TheBLG-enriched composition comprises BLG in an amount of at least 85% w/wrelative to total protein, and preferably at least 90% w/w. In somecases the BLG-enriched composition can be used directly as the liquidBLG isolate. However, often additional processing is required to convertthe BLG-enriched composition to the liquid BLG isolate.

In the context of the present invention, the term “whey proteinsolution” is used to describe the special aqueous whey proteincomposition that is supersaturated with respect to BLG in salting-inmode and useful for preparing BLG crystals.

In the context of the present invention, the term “sterile” means thatthe sterile composition or product in question does not contain anyviable microorganisms and therefore is devoid of microbial growth duringstorage at room temperature. A composition that has been sterilized issterile.

When a liquid, such as a beverage preparation, is sterilized andpackaged aseptically in a sterile container it typically has a shelflife of at least six months at room temperature. The sterilizationtreatment kills spores and microorganisms that could cause spoilage ofthe liquid.

In the context of the present invention the term “energy content” meansthe total content of energy contained in a food product. The energycontent can be measured in kilojoule (kJ) or kilo calories (kcal) andare referred to as calories per amount of food product, e.g. kcal per100 grams of the food product. One example is an instant beverage powderhaving an energy content of 350 kcal/100 grams of the instant beveragepowder.

The total energy content of a food product includes the energycontribution from all the macronutrients present in the food product,e.g. energy from protein, lipid and carbohydrate. The distribution ofenergy from the macronutrients in the food product can be calculatedbased on the amount of the macronutrients in the food product and thecontribution of the macronutrient to the total energy content of thefood product. The energy distribution can be stated as energy percent (E%) of the total energy content of the food product. For example for aninstant beverage powder comprising 20 E % protein, 50 E % carbohydrateand 30 E % lipid, this means that 20% of the total energy comes fromprotein, 50% of the total energy comes from carbohydrate and 30% of thetotal energy comes from fat (lipid).

In the context of the present invention the term “nutritionalsupplement” pertains to a food product comprising one or more macronutrients such as protein, lipid and/or carbohydrate and optionallycomprising vitamins and minerals. Nutritional supplements can be eithercomplete or incomplete.

By the term “nutritionally complete nutritional supplement” isunderstood food products comprising protein, lipid and carbohydrate andfurther comprising vitamins, minerals and trace elements, where the foodproduct has a nutrient profile matching a complete and healthy diet.

The term “nutritionally incomplete supplement” means food productscomprising one or more macro nutrients and optionally further comprisingvitamins, minerals and trace elements. A incomplete nutritionallysupplement may comprise protein as the only nutrients or may compriseprotein, lipid and a carbohydrate.

The term “food for special medical purposes (FSMP)” or “medical food”are food products for oral ingestion or tube feeding, which are used forspecific medical disorders, diseases or conditions for which there aredistinctive nutritional requirements and which are used under medicalsupervision. A medical food can be a nutritionally complete supplementor a nutritionally incomplete supplement.

The term “nutrient” means a substance used by an organism to survive,grow and reproduce. Nutrients can be either macronutrients ormicronutrients. Macronutrients are nutrients that provide energy whenconsumed e.g. protein, lipid and carbohydrate. Micronutrients arenutrients are vitamins, minerals and trace elements.

By the term “instant beverage powder” or “instant beverage powderproduct” is meant a powder which can be converted to a liquid beverageby addition of a liquid, such as water.

In the context of the present invention the terms “beverage preparation”and “preparation” used as a substantive relate to any water-based liquidwhich can be ingested as a drink, e.g. by pouring, sipping ortube-feeding.

In the context of the present invention the term “protein fraction”relates to proteins of the composition in question e.g. the proteins ofa powder or a beverage preparation.

In the context of the present invention the term “astringency” relatesto a mouthfeeling. Astringency feels like a contraction of cheek musclesand results in increased saliva production. Thus, astringency is not ataste as such, but a physical mouthfeeling and time-dependent feeling inthe mouth.

In the context of the present invention the term “drying mouthfeeling”relates to a feeling in the mouth, it feels like a drying of the mouthand teeth and results in minimization of the saliva production. Thusdrying mouthfeeling is not a taste as such, but a physical mouthfeelingand time-dependent feeling in the mouth.

In the context of the present invention the term “minerals” as usedherein, unless otherwise specified, refers to any one of major minerals,trace or minor minerals, other minerals, and combinations thereof. Majorminerals include calcium, phosphorus, potassium, sulfur, sodium,chlorine, magnesium. Trace or minor minerals include iron, cobalt,copper, zinc, molybdenum, iodine, selenium, manganese and other mineralsinclude chromium, fluorine, boron, lithium, and strontium.

In the context of the present invention the terms “lipid”, “fat”, and“oil” as used herein unless otherwise specified, are usedinterchangeably to refer to lipid materials derived or processed fromplants or animals. These terms also include synthetic lipid materials solong as such synthetic materials are suitable for human consumption.

In the context of the present invention the term “transparent”encompasses a beverage preparation having a visibly clear appearance andwhich allows light to pass and through which distinct images appear. Atransparent beverage has a turbidity of at most 200 NTU.

In the context of the present invention the terms “opaque” encompasses abeverage preparation having a visibly unclear appearance and it has aturbidity of more than 200 NTU.

In the context of the present invention the term “mother liquor”pertains to the whey protein solution that remains after BLG has beencrystallised and the BLG crystals have be at least partially removed.The mother liquor may still contain some BLG crystals but normally onlysmall BLG crystals that have escaped the separation.

By the term “instant beverage powder” or “instant beverage powderproduct” is meant a powder which can be converted to a liquid beverageby addition of a liquid, such as water.

DETAILED DESCRIPTION

The overall conception of the nutritional supplement is noticed by theconsumer. The nutritional supplement should be appetizing in taste andappearance; otherwise it will be rejected by the consumer. Further, theconsumer values natural products without additives. A further parameterof importance to the consumer is shelf-life of the product.

An aspect of the invention pertains to an instant beverage powdercomprising at least 1% w/w BLG, preferably at least 5%, wherein:

-   -   i. the crystallinity of BLG is at least 20%, preferably at least        40%, and/or    -   ii. at least 85% w/w of the total amount of protein is comprised        by BLG,        and furthermore comprising at least one additional ingredient        selected from the group consisting of vitamins, flavouring        agent, colouring agent, minerals, sweeteners, antioxidants, food        acid, lipids, carbohydrate, prebiotics, probiotics, anti-foaming        agents and non-whey protein.

The BLG source used in the instant beverage powder can be the BLGisolate or BLG isolate powder as described in the present patentapplication. In some preferred embodiments of the invention the BLGsource contributes with at least 90% w/w of the total protein of theinstant beverage powder, more preferably at least 95% w/w, even morepreferred at least 98% w/w and most preferred all the protein of theinstant beverage powder.

In some preferred embodiments of the invention the BLG source is the BLGisolate powder and it is the only source of protein in the instantbeverage powder.

In some preferred embodiments of the invention the instant beveragepowder is prepared by dry-blending the BLG isolate powder and the otheringredients.

In other preferred embodiments of the invention the instant beveragepowder is prepared by using at least one ingredient in dissolved formand subsequently performing a drying steps. The drying step may e.g.form part of a wet-granulation process or a spray-drying step.

The BLG of the instant beverage powder of the present inventionpreferably has a low degree of denaturation, such as at most 10%,preferably at most 4%, more preferably at most 1%, even more preferablyat most 0.4% and even more preferably at most 0.1%. Most preferably, theBLG is not denatured at all. For instant beverage powders it isadvantageous that BLG has a low degree of denaturation, as this reducesthe tendency to foam when mixed with a liquid.

The instant beverage powder of preferably has a degree of proteindenaturation of at most 10%, preferably at most 4%, more preferably atmost 1%, even more preferably at most 0.4% and even more preferably atmost 0.1%. Most preferably, the protein is not denatured at all.

In one embodiment of the invention the instant beverage powder comprisesfrom 1-90% w/w BLG. In a preferred embodiment of the invention, theinstant beverage powder comprises from 30-90% w/w BLG, more preferablyin the range of 40-90% w/w BLG or even more preferably in the range of50-90% w/w BLG.

In other preferred embodiments of the invention the instant beveragepowder comprises from 10-97% w/w BLG. In a preferred embodiment of theinvention, the instant beverage powder comprises from 30-96% w/w BLG,more preferably in the range of 40-95% w/w BLG or even more preferablyin the range of 50-94% w/w BLG.

In one embodiment of the invention, the instant beverage powdercomprises from 1-50% w/w BLG. In a preferred embodiment of theinvention, the instant beverage powder comprises from 2-45% w/w BLG,more preferably in the range of 3-40% w/w BLG or even more preferably inthe range of 3-35% w/w BLG.

In a preferred embodiment of the invention, the instant beverage powdercomprises at least 85% w/w of the total amount of protein is BLG.

In one embodiment of the invention, the instant beverage powdercomprises at least 85% w/w BLG relative to total protein such as atleast 86% w/w BLG relative to total protein, at least 87% w/w BLGrelative to total protein, at least 88% w/w BLG relative to totalprotein, at least 89% w/w BLG relative to total protein.

In one embodiment of the invention, the instant beverage powdercomprises at least 91% w/w BLG relative to total protein such as atleast 92% w/w BLG relative to total protein, at least 93% w/w BLGrelative to total protein, at least 94% w/w BLG relative to totalprotein, at least 95% w/w BLG relative to total protein, at least 96%w/w BLG relative to total protein, at least 97% w/w BLG relative tototal protein, at least 98% w/w BLG relative to total protein or atleast 99% w/w BLG relative to total protein.

In some preferred embodiments of the instant beverage powder of theinvention, at least 85% w/w of the protein is BLG. Preferably, at least88% w/w of the protein is BLG, more preferably at least 90% w/w, evenmore preferably at least 91% w/w, and most preferably at least 92% w/wof the protein is BLG.

Even higher relative amounts of BLG are both feasible and desirable thusin some preferred embodiments of the invention at least 94% w/w of theprotein of the instant beverage powder is BLG, more preferably at least96% w/w of the protein is BLG, even more preferably at least 98% w/w ofthe protein is BLG, and most preferably approx. 100% w/w of the proteinis BLG.

For example, the instant beverage powder preferably comprises BLG in anamount of at least 97.5% w/w relative to total protein, preferably atleast 98.0% w/w, more preferably at least 98.5% w/w, even morepreferably at least 99.0%, and most preferably BLG in an amount of atleast 99.5% w/w relative to total protein, such as approx. 100.0% w/wrelative to total protein.

The protein of the instant beverage powder is preferably prepared frommammal milk, and preferably from ruminant milk such as e.g. milk fromcow, sheep, goat, buffalo, camel, llama, horse and/or deer. Proteinderived from bovine milk is particularly preferred. The protein of theinstant beverage powder is therefore preferably bovine milk protein.

The protein of the instant beverage powder is preferably whey protein ormilk serum protein and even more preferably bovine whey protein or milkserum protein.

The intrinsic tryptophan fluorescence emission ratio (I330 nm/I350 nm)is a measure of the degree of unfolding of BLG, and the inventors havefound that at high BLG tryptophan fluorescence emission ratios, whichcorrelate with low or no unfolding of BLG, the intrinsic tryptophanfluorescence emission ratio (I330 nm/I350 nm) is measured according toExample 1.1.

In some preferred embodiments of the invention, the instant beveragepowder has an intrinsic tryptophan fluorescence emission ratio (I330nm/I350 nm) of at least 1.11.

In some preferred embodiments of the invention, the instant beveragepowder has an intrinsic tryptophan fluorescence emission ratio (I330nm/I350 nm) of at least 1.12, preferably at least 1.13, more preferablyat least 1.15, even more preferably at least 1.17, and most preferablyat least 1.19.

If instant beverage powder contains considerable amounts of non-proteinmatter, it is preferred to isolate the protein fraction before measuringthe intrinsic tryptophan fluorescence emission ratio. Thus in somepreferred embodiments of the invention, the protein fraction of instantbeverage powder has an intrinsic tryptophan fluorescence emission ratioof at least 1.11.

In some preferred embodiments of the invention, the protein fraction ofthe instant beverage powder has an intrinsic tryptophan fluorescenceemission ratio (I330 nm/I350 nm) of at least 1.12, preferably at least1.13, more preferably at least 1.15, even more preferably at least 1.17,and most preferably at least 1.19.

The protein fraction can e.g. be separated from the instant beveragepowder by dissolving the instant beverage powder in demineralised waterand subjecting the solution to dialysis or ultrafiltration-baseddiafiltration using a filter that retains the protein.

In some preferred embodiments of the invention the crystallinity of BLGof the instant beverage powder is at least 20%, preferably at least 40%,more preferably at least 60%, even more preferably at least 80%, andmost preferably at least 90%. Having a crystallinity of BLG of at least20% means that a significant amount of the BLG is present in the form ofdried BLG crystals in the instant beverage powder.

The present inventors have found that a crystallinity of BLG of at least20% is advantageous as it means that the protein is present in a formthat has a higher density than traditional WPI. This provides a higheroverall bulk density to the instant beverage powder and makes it lessdusty and easier to handle for the end user. The inventors have alsoobserved a reduced tendency to particle segregation in dry-blendedinstant beverage powders that e.g. contain a carbohydrate powder and/orfood acid powder in addition to a protein powder.

The present invention makes it possible to provide low carbohydrateinstant beverage powder which have both sweetness and a high proteincontent.

Thus, in some preferred embodiments of the invention the instantbeverage powder comprises:

-   -   an energy content in the range of 320-380 kcal/100 grams of        powder, and preferably in the range of 350-370 kcal/100 grams,    -   a contribution of the energy from protein in the range of 90-100        E %, and preferably in the range of 95-100 E %,    -   BLG in an amount of at least 85% w/w relative to total protein,        preferably at least 90% w/w relative to total protein, and more        preferably at least 94% w/w relative to total protein    -   a contribution of the energy from carbohydrate in the range of        0-10 E %, and preferably in the range of 0-5 E %,    -   a total amount of high intensity sweeteners in the range of        0.01-4% w/w, preferably in the range of 0.05-3%    -   having a bulk density of at least 0.45 g/mL, preferably at least        0.50 g/mL, and more preferred at least 0.6 g/mL.

The protein of the instant beverage powder is preferably provided by aBLG isolate powder that: has a pH in the range of i) 2-4.9, ii) 6.1-8.5,or iii) 5.0-6.0 and comprises:

-   -   total protein in an amount of at least 90% w/w, preferably at        least 95% w/w,    -   BLG in an amount of at least 85% w/w relative to total protein,        preferably at least 90% w/w relative to total protein, and more        preferably at least 94% w/w relative to total protein, said BLG        isolate powder having:    -   a bulk density of at least 0.45 g/mL, preferably at least 0.50        g/mL, and more preferred at least 0.6 g/mL, and    -   one or more of the following:        -   an intrinsic tryptophan fluorescence emission ratio            (I330/I350) of at least 1.11, preferably at least 1.13, and            more preferably at least 1.15        -   a degree of protein denaturation of at most 10%, preferably            at least 5%        -   a heat-stability at pH 3.9 of at most 200 NTU, and        -   at most 1000 colony-forming units/g.

In one embodiment of the invention the instant beverage powder furthercomprises at least one additional ingredient selected from the groupconsisting of vitamins, flavouring agent, colouring agent, minerals,sweeteners, antioxidants, food acid, lipids, carbohydrate, prebiotics,probiotics and non-whey protein.

The further ingredient ensures that the instant beverage powder containsthe desired nutrients, i.e. nutrients specifically adapted to a patientswith or at risk of malnutrition, for patients suffering from kidneydisease, for weight gain or it can be used as a nutritional supplement,e.g. by sportsmen or athletes.

In a preferred embodiment of the invention, the instant beverage powdermay include a vitamin selected from the group consisting of vitamin A,vitamin B1 (thiamine), vitamin B2 (riboflavin), vitamin B3 (niacin),vitamin B5 (pantothenic acid), vitamin B6 (pyridoxine), vitamin B7(biotin), vitamin B9 (folic acid), and vitamin B12 (cobalamine), vitaminC, vitamin D, vitamin E, vitamin K, choline, inositol, their salts,their derivatives and combinations thereof.

In an embodiment, the instant beverage powder may comprise a flavouringagent selected from the group consisting of salt, flavorings, flavorenhancers and/or spices. In a preferred embodiment of the invention theflavor comprise chocolate, cocoa, lemon, orange, lime, strawberry,banana, forrest fruit flavor or combinations thereof.

In an embodiment, the instant beverage powder may include a mineralselected from the group consisting of boron, calcium, chromium, copper,iodine, iron, magnesium, manganese, molybdenum, nickel, phosphorus,potassium, sodium, selenium, silicon, tin, vanadium, zinc, orcombinations thereof.

The instant beverage powder may furthermore contain salts and mineralswhich typically are present in whey or milk derived products. Themineral content of instant beverage powder are typically represented asthe ash content of the food ingredient or product. In a preferredembodiment, the instant beverage powder may comprise an antioxidantselected from the group consisting of beta-carotene, vitamin C, vitaminE, selenium, or combinations thereof.

In an embodiment of the invention, the instant beverage powder maycomprise one or more sweeteners, such as carbohydrate sweeteners,polyols and/or high intensity sweeteners. The instant beverage powdermay e.g. comprise a total amount of carbohydrate sweetener in the rangeof 0.001-20% w/w relative to the total weight of the instant beveragepowder. Alternatively, the instant beverage powder may comprise a totalamount of carbohydrate sweetener in the range of 0.1-15% w/w relative tothe total weight of the food product.

In one embodiment of the invention, the instant beverage powdercomprises at least one high intensity sweetener. In one embodiment, theat least one high intensity sweetener is selected from the groupconsisting of aspartame, cyclamate, sucralose, acesulfame salt, neotame,saccharin, stevia extract, a steviol glycoside such as e.g. rebaudiosideA, or a combination thereof. In some embodiments of the invention, it isparticularly preferred that the sweetener comprises or even consists ofone or more high intensity sweeteners (HIS).

High intensity sweeteners are both found among both natural andartificial sweeteners and typically have a sweetening intensity of atleast 10 times that of sucrose.

If used, the total amount of high intensity sweeteners is typically inthe range of 0.01-4% w/w. For example, the total amount of highintensity sweeteners may be in the range of 0.05-3% w/w. Alternatively,the total amount of high intensity sweeteners may be in the range of0.1-2.0% w/w.

The choice of the sweetener may depend on the beverage to be produced,and the consumer of the product, e.g. it may be adjusted to a specificdiagnosis of a patient. High-intensity sugar sweeteners (e.g. aspartame,acetsulfam-K or sucralose) may be used in beverage where no energycontribution from the sweetener is desired, whereas for beverages havinga natural profile natural sweeteners (e.g. steviol glycosides, sorbitolor sucrose) may be used.

It may furthermore be preferred that the sweetener comprises or evenconsists of one or more polyol sweetener(s). Non-limiting examples ofuseful polyol sweetener are maltitol, mannitol, lactitol, sorbitol,inositol, xylitol, threitol, galactitol or combinations thereof. Ifused, the total amount of polyol sweetener is typically in the range of1-40% w/w. For example, the total amount of polyol sweetener may be inthe range of 2-30% w/w. Alternatively, the total amount of polyolsweetener may be in the range of 4-20% w/w.

In one embodiment of the invention the instant beverage powder maycomprise one or more of:

-   -   i. a sweetener, e.g. a sugar sweetener and/or a non-sugar        sweetener,    -   ii. a flavoring agent,    -   iii. at least one food acid, e.g. citric acid or other suitable        food acids,    -   iv. the sum of the amounts of Na, K, Mg, and Ca of the instant        beverage is at most 10 mmol/g protein and        wherein a 10% w/w solution of the powder in demineralized water        has a pH in the range of 2-8.

The pH of the instant beverage powder can be measured by dissolving 10gram of the instant beverage powder in 90 ml of demineralized water atroom temperature, as described in example 1.16.

The inventors have found that it is advantageous to use a lowphosphorus/low potassium BLG isolate powder in the instant beveragepowder, e.g. for instant beverage powders that are particularly usefulto patients with kidney diseases.

By adding sweetener, flavoring agents and/or food acids, the taste ofthe product can be designed so that the instant beverage powder isappealing to the consumer. In one embodiment of the invention theconsumer can be a patient for which the flavor, sweetener and acidicprofile of the instant beverage powder is adjusted to fit to thepatients need and diagnosis.

In a preferred embodiment of the invention the instant beverage powdermay comprise a high intensity sweetener and a flavoring agent. In aneven more preferred embodiment of the invention, the instant beveragepowder comprises 0.001-0.05% w/w sucralose and 0.01-0.2% w/w and aflavor selected from chocolate, cocoa, lemon, orange, lime, strawberry,banana, forrest fruit flavor or combinations thereof.

In one embodiment of the invention, the instant beverage powdercomprises an anti-foaming agent. The anti-foaming agent may be selectedfrom anti-foaming agents suitable for food products. The anti-foamingagent may be selected from oil-based anti-foaming agents, water-basedanti-foaming agents, silicone-based anti-foaming agents, EP/PO-basedanti-foaming agents or a combination thereof.

The instant beverage powder has a water content of at most 6% w/w. Inone embodiment of the invention, the instant beverage powder comprisesat most 5% w/w water, preferably at most 4% w/w water, more preferablyat most 3% w/w water, and even more preferably at most 2% w/w water.

The storage stability of the instant beverage powder may increase whenlowering the water content of the powder.

The present inventors have found that it can be advantageous to controlthe mineral content in order to reach some of the desired properties ofthe instant beverage powder.

In some preferred embodiments of the invention, the sum of the amountsof Na, K, Mg, and Ca of the instant beverage powder is at most 10 mmol/gprotein. Preferably, the sum of the amounts of Na, K, Mg, and Ca of theinstant beverage powder is at most 6 mmol/g protein, more preferably atmost 4 mmol/g protein, even more preferably at most 2 mmol/g protein.

In other preferred embodiments of the invention the sum of the amountsof Na, K, Mg, and Ca of the instant beverage powder is at most 1 mmol/gprotein. Preferably, the sum of the amounts of Na, K, Mg, and Ca of theinstant beverage powder is at most 0.6 mmol/g protein, more preferablyat most 0.4 mmol/g protein, even more preferably at most 0.2 mmol/gprotein, and most preferably at most 0.1 mmol/g protein.

In one embodiment of the invention, the instant beverage powdercomprises dry BLG crystals, e.g. obtainable by one or more methodsdescribed in PCT/EP2017/084553. The instant beverage powder containingBLG crystals may have a bulk density of at least 0.30 g/mL, preferablyat least 0.4 g/mL.

The inventors have observed that instant beverage powder in which atleast some of BLG is on crystal form has a higher density thancomparable instant BLG compositions without BLG crystals. Thus, in somepreferred embodiments of the invention the instant beverage powder has abulk density of at least 0.30 g/mL, preferably at least 0.40 g/mL.Preferably the instant beverage powder has a bulk density of at least0.45 g/mL. More preferably the instant beverage powder has a bulkdensity of at least 0.50 g/mL. It is even more preferred that theinstant beverage powder has a bulk density of at least 0.6 g/mL. Theinstant beverage powder may e.g. have a bulk density of at least 0.7g/mL.

The instant beverage powder of the present invention preferably has abulk density in the range of 0.3-1.0 g/mL, preferably in the range of0.40-0.9 g/mL, more preferably in the range of 0.45-0.8 g/mL, even morepreferably in the range of 0.45-0.75 g/mL, even more preferably in therange of 0.50-0.75 g/mL, and most preferably in the range of 0.6-0.75g/mL.

The bulk density of a powder can be measured according to Example 1.17.

The total protein content and the energy content in the instant beveragepowder of the invention depend on the intended use of the instantbeverage powder. The energy content of an instant beverage powder is inthe range of 200-500 kcal/100 grams of powder.

For instant beverage powders, the contribution of the energy fromprotein may be at least 7 E %, preferably at least 25 E %, morepreferably at least 30 E %, even more preferably at least 40 E %.

In a preferred embodiment of the invention the contribution of energyfrom protein is in the range of 10-30 E %, preferably in the range of10-15 E % or even more preferably 11 E %. Alternatively, thecontribution of energy from protein is in the range of 15-25 E %,preferably in the range of 18-22 E %.

In a preferred embodiment of the invention, the contribution of theenergy from protein is in the range of 7-25 E %, preferably in the rangeof 10-25 E %, more preferably 15-20 E % or even more preferably theinstant beverage powder contains 15 E % from protein or 20 E % fromprotein, or the contribution of the energy from protein is in the rangeof 8-15 E %.

In another preferred embodiment of the invention the contribution of theenergy from protein is at least 50 E %, preferably at least 60 E % or atleast 70 E % or even more preferred at least 80 E %. In a preferredembodiment of the invention, the contribution of the energy from proteinis in the range of 80-100 E %, preferably in the range of 90-100 E % oreven more preferably in the range of 95-100 E %.

In a preferred embodiment of the invention, the contribution of theenergy from protein is in the range of 30-80 E %, preferably in therange of 60-80 E %. In another embodiment of the invention thecontribution of the energy from protein is in the range of 30-40 E % oreven more preferably the instant beverage powder contains 33 E % fromprotein.

The instant beverage powder can be used as a nutritional supplement e.g.for treating patients with or at risk of malnutrition, for patientssuffering from kidney disease, for weight gain or can be used as anutritional supplement e.g. by sportsmen or athletes, before, during orafter exercise.

The instant beverage powder of the present invention may comprise othermacronutrients than protein. The instant beverage powder can comprisecarbohydrates and/or lipids in addition to the protein. The total lipidcontent in the instant beverage powder of the invention depends on theintended use of the instant beverage powder. In one embodiment of theinvention the contribution of energy from lipid is in the range of 0-60E %

In a preferred embodiment of the invention the contribution of energyfrom lipid is in the range of 0-5 E %, preferably in the range of 0-3 E% or more preferred in the range of 0-2 E % from lipid.

Even less lipid may be preferred, thus in a preferred embodiment of theinvention the contribution of energy from lipid is in the range of 0-1 E%, preferably in the range of 0-0.1 E % or more preferred in the rangeof 0-0.01 E % from lipid.

In a preferred embodiment of the invention the contribution of theenergy from lipid is in the range of 30-60 E %, preferably in the rangeof 30-50 E % or even more preferably the instant beverage powdercontains 35 E % from lipid, 45 E % from lipid or 50 E % from lipid.Alternatively the contribution of the energy from lipid is in the rangeof 25-45 E %.

In a preferred embodiment of the invention, the contribution of theenergy from lipid is in the range of 15-20 E %, preferably in the rangeof 16-18 E % or even more preferably the instant beverage powdercontains 16 E % from lipid.

The instant beverage powder can be used as a nutritional supplement e.g.for treating patients with or at risk of malnutrition, for patientssuffering from kidney disease, for weight gain or can be used as anutritional supplement e.g. by sportsmen or athletes, before, during orafter exercise.

In one embodiment of the invention, the instant beverage powder maycomprise a carbohydrate in addition to protein. The energy contributionof the carbohydrate to the total energy of the instant beverage powdermay be in the range of 0-90 E %.

The carbohydrate can be selected from sugars, oligosaccharides orpolysaccharides. Examples of sugars are mono-, disaccharides andpolyols. Examples of oligosaccharides are malto-oligosaccharides, suchas maltodextrins or other oligosaccharides, such as raffinose,stachyouse or fructo-oligosaccharides. Examples of polysaccharides arestarches such as amylose, amylopectins, or modified starches andnon-starch polysaccharides, such as dietary fibers, cellulose, pectinsand hydrocolloids. In a preferred embodiment of the invention, thecarbohydrate is selected from maltodextrine, saccharose or glucosesyrup.

The total carbohydrate content in the instant beverage powder of theinvention depends on the intended use of the instant beverage powder.For instant beverage powders used for sportsmen or athletes,carbohydrate in the form of sugars may be added in order to boostimmediate energy for the sportsman or carbohydrate in the form of slowcarbohydrates or dietary fiber may be added in order to prolong satiety.

In a preferred embodiment of the invention, the contribution of energyfrom carbohydrate is in the range of 70-90 E %, preferably in the rangeof 75-85 E % or more preferably the instant beverage powder contains 89E % from carbohydrate

In a preferred embodiment of the invention, the contribution of theenergy from carbohydrate is in the range of 30-50 E %, preferably in therange of 35-45 E % or even more preferably the instant beverage powdercontains 35 E % from carbohydrate, 45 E % from carbohydrate or 50 E %from carbohydrate. Alternatively, the contribution of the energy fromcarbohydrate is in the range of 40-60 E %, such as in the range of 45-55E %.

In another preferred embodiment of the invention, the contribution ofthe energy from carbohydrate is in the range of 0-20 E %. In a preferredembodiment of the invention the contribution of the energy fromcarbohydrate is in the range of 0-10 E %, preferably in the range of 0-5E %.

In yet a preferred embodiment of the invention, the contribution of theenergy from carbohydrate is in the range of 0-4 E %, more preferably 0-1E %, and even more preferably 0-0.2 E %.

In another preferred embodiment of the invention the contribution of theenergy from carbohydrate is in the range of 3-20 E %. In a preferredembodiment of the invention, the contribution of the energy fromcarbohydrate is in the range of 4-15 E %. In another embodiment of theinvention the contribution of the energy from carbohydrate is in therange of 45-55 E %.

The instant beverage powder can be used as a nutritional supplement e.g.for treating patients with or at risk of malnutrition, for patientssuffering from kidney disease, for weight gain or can be used as anutritional supplement e.g. by sportsmen or athletes, before, during orafter exercise.

The instant beverage powder of the present invention comprises proteinand may in addition to the protein comprise lipid and/or carbohydratedepending on the intended use of the instant beverage powder.

In a preferred embodiment of the invention, the instant powder furthercomprises vitamins, minerals and trace elements. The instant beveragepowder can be used as a nutritional supplement e.g. for treatingpatients with or at risk of malnutrition, for patients suffering fromkidney disease, for weight gain or it can be used as a nutritionalsupplement e.g. by sportsmen or athletes.

The energy content of an instant beverage powder can be in the range of200-400 kcal/100 grams of powder. In a preferred embodiment of theinvention, the energy content of the instant beverage powder is in therange of 300-400 kcal/100 grams of powder, even more preferred theenergy content of the instant beverage powder is in the range of 320-380kcal/100 grams of powder, or most preferred in the range of 350-370kcal/100 grams of powder.

In a preferred embodiment of the invention, the instant beveragesupplement has an energy distribution as follows 10-30 E % protein,70-90 E % carbohydrates and 0-5 E % lipid. In a more preferredembodiment of the invention the instant beverage supplement has anenergy distribution of 10-15 E % protein, 75-85 E % carbohydrates and0-1 E % lipid. In a preferred embodiment of the invention the instantbeverage supplement has an energy distribution of 11 E % protein, 89 E %carbohydrates and 0 E % lipid.

In one embodiment of the invention the instant beverage powder comprisesprotein, carbohydrate and lipid and optionally comprising vitamins,minerals and trace elements. The instant beverage powder can be designedso that the recommended daily intake of the beverage powder supplies therecommended daily intake of the vitamins, minerals and trace elements.However this is not a requirement.

Such instant beverage powder may be useful as nutritional supplementwhere the consumer is interested in a nutritional supplement with aproportion of macronutrients reflecting a healthy diet with respect toenergy distribution, macronutrients and micronutrients, e.g. where thenutritional supplement is given under supervision of a health careprofessional.

The energy content of an instant beverage powder is in the range of400-500 kcal/100 grams of powder. In a preferred embodiment of theinvention, the energy content of the instant beverage powder is in therange of 410-490 kcal/100 grams of powder, even more preferably theenergy content of the instant beverage powder is in the range of 420-480kcal/100 grams of powder or most preferred in the range of 440-460kcal/100 grams of powder.

In a preferred embodiment of the invention, the instant beveragesupplement has an energy distribution as follows; 7-25 E % protein,30-50 E % carbohydrates and 30-55 E % lipid. In a more preferredembodiment of the invention the instant beverage supplement has anenergy distribution of 10-25 E % protein, 30-50 E % carbohydrates and30-55 E % lipid, or even more preferably 15-20 E % protein, 35-45 E %carbohydrates and 35-50 E % lipid. In an even more preferred embodimentof the invention the instant beverage supplement has an energydistribution of 15 E % protein, 35 E % carbohydrates and 50 E % lipid,has an energy distribution of 20 E % protein, 45 E % carbohydrates and35 E % lipid or has an energy distribution of 8-15 E % protein, 40-47 E% carbohydrates and 45 E % lipid.

In a preferred embodiment of the invention, the instant powder furthercomprises vitamins, minerals and trace elements.

The instant beverage powder can be used as a nutritional supplement e.g.for treating patients with or at risk of malnutrition, for patientssuffering from kidney disease, for weight gain or can be used as anutritional supplement e.g. by sportsmen or athletes, before, during orafter exercise.

In one embodiment of the invention, the instant beverage powdercomprises protein, carbohydrate and lipid. Such instant beverage powdermay be useful as nutritional supplement where intake of protein is ofhighest priority of the consumer, e.g. where the consumer would like tosupplement the regular meals. The energy content of an instant beveragepowder is in the range of 200-500 kcal/100 grams of powder. In apreferred embodiment of the invention, the energy content of the instantbeverage powder is in the range of 200-350 kcal/100 grams of powder, oreven more preferably the energy content of the instant beverage powderis in the range of 200-300 kcal/100 grams of powder.

In a preferred embodiment of the invention, the instant beveragesupplement has an energy distribution as follows 80-98 E % protein, 0-20E % carbohydrates and 0-5 E % lipid. In a more preferred embodiment ofthe invention the instant beverage supplement has an energy distributionof 90-98 E % protein, 0-10 E % carbohydrates and 0-3 E % lipid. In aneven more preferred embodiment of the invention the instant beveragesupplement has an energy distribution of 95-98 E % protein, 0-5 E %carbohydrates and 0-2 E % lipid.

In an embodiment of the invention, the instant powder further comprisesvitamins, minerals and trace elements. In a preferred embodiment of theinvention, the instant powder further comprises vitamins, minerals andtrace elements. The instant beverage powder can be used as a nutritionalsupplement e.g. for treating patients with or at risk of malnutrition,for patients suffering from kidney disease, for weight gain or can beused as a nutritional supplement e.g. by sportsmen or athletes.

In one embodiment of the invention the instant beverage powder comprisesprotein, carbohydrate and lipid. The energy content of an instantbeverage powder is in the range of 200-420 kcal/100 grams of powder. Ina preferred embodiment of the invention, the energy content of theinstant beverage powder is in the range of 300-420 kcal/100 grams ofpowder, or more preferred the energy content of the instant beveragepowder is in the range of 320-380 kcal/100 grams of powder, or even morepreferred the energy content of the instant beverage powder is in therange of 350-370 kcal/100 grams of powder.

In a preferred embodiment of the invention, the instant beveragesupplement has an energy distribution as follows; 30-80 E % protein,3-20 E % carbohydrates and 15-20 E % lipid. In a more preferredembodiment of the invention, the instant beverage supplement has anenergy distribution of 60-80 E % protein, 4-15 E % carbohydrates and16-18 E % lipid. In an even more preferred embodiment of the inventionthe instant beverage supplement has an energy distribution of 30-40 E %protein, 45-55 E % carbohydrates and 12-18 E % lipid, such as eg. 33 E %protein, 46 E % carbohydrates and 15 E % lipid.

Alternatively, the energy content of an instant beverage powder can bein the range of 150-250 kcal/100 grams of powder with an energydistribution as follows; 10-30 E % protein, 40-60 E % carbohydrates and25-45 E % lipid, or preferably 15-25 E % protein, 45-55 E %carbohydrates and 30-40 E % lipid.

In an embodiment of the invention, the instant powder further comprisesvitamins, minerals and trace elements.

In a preferred embodiment of the invention, the instant powder furthercomprises vitamins, minerals and trace elements. The instant beveragepowder can be used as a nutritional supplement e.g. for treatingpatients with or at risk of malnutrition, for patients suffering fromkidney disease, for weight gain or it can be used as a nutritionalsupplement e.g. by sportsmen or athletes.

In a preferred embodiment of the invention, the instant beverage powdermay further comprise a flavouring agent, colouring agent, sweeteners,antioxidants, food acid, lipids, carbohydrate, prebiotics, probiotics ornon-whey protein.

For reasons of convenience the instant beverage powder may be sold in akit comprising the instant powder of the invention, a tool for measuringsaid powder, and a container having a lid for opening and closing thecontainer, wherein said container is for mixing said powder with aliquid to form a food product, and said container is adapted fordrinking the food product directly from the container. Examples ofuseful containers are e.g. bottles, cartons, bricks, pouches and/orbags.

The consumer buying the kit will obtain all items for readily preparinga liquid food product according to the invention. The measuring toolensures that the consumer weights out the correct amount of instantpowder for the amount of water in the container.

In one embodiment of the invention, the tool for measuring the instantpowder is a spoon and the container is a drinking bottle. In oneembodiment of the invention the container has an inside indication ofhow much liquid to fill in the container. In one embodiment of theinvention the lid has an opening adapted for drinking the liquid foodproduct directly from the container and for closing while mixing theliquid food product.

The pH of the instant beverage powder is important because the taste ofthe product prepared from the instant beverage powder depend on the pHof the product. The pH of the powder can be determined by measuring thepH in a 10% w/w solution of the instant beverage powder in demineralisedwater at 25° C., as described in example 1.16. In one embodiment of theinvention, the pH of the instant beverage powder in a 10% w/w solutionin demineralised water is in the range of 2-8 at 25° C.

In one embodiment of the invention, the pH is in the range of 2.0-4.9,such as in the range of in the range of 2.5-4.7, more preferably2.8-4.3, even more preferably 3.2-4.0, and most preferably 3.4-3.9.Alternatively, but also preferred, the instant beverage powder may havea pH in the range of 3.6-4.3.

Alternatively, the pH of the instant beverage powder in a 10% w/wsolution in demineralised water is a pH in the range of 5.0-6.0 at 25°C., preferably, the powder has a pH in the range of 5.1-5.9, morepreferably 5.2-5.8, even more preferably 5.3-5.7, and most preferably5.4-5.6.

Alternatively, the pH of the instant beverage powder is in the range of6.1-8.5, more preferably 6.2-8.0, even more preferably 6.3-7.7, and mostpreferably 6.5-7.5.

Yet an aspect of the invention pertains to the use of the instantbeverage as defined herein as a food ingredient.

An aspect of the invention relates to a packaged instant beverage powderproduct comprising a container containing the instant beverage powderproduct as described herein.

In an embodiment of the invention, the instant beverage powder productis hermetically sealed in the container, optionally packaged with aninert gas.

A wide range of different containers may be used to store the instantbeverage powder product. For example, the container may be a containerselected from the group consisting of a bottle, a can, a bag, a pouch,and a sachet.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-500 kcal/100 grams of powder and theinstant beverage powder comprises at least 85% w/w BLG relative to totalprotein, preferably at least 90% w/w BLG relative to total protein.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-500 kcal/100 grams of powder and theinstant beverage powder comprises at least 85% w/w BLG relative to totalprotein, preferably at least 90% w/w BLG relative to total protein andthe pH of the powder is in the range of 2-8.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-500 kcal/100 grams of powder and theinstant beverage powder comprises at least 85% w/w BLG relative to totalprotein, preferably at least 90% w/w BLG relative to total protein andthe pH of the powder is in the range of 2.0-4.9.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-500 kcal/100 grams of powder and theinstant beverage powder comprises at least 85% w/w BLG relative to totalprotein, preferably at least 90% w/w BLG relative to total protein andthe pH of the powder is in the range of 5.0-6.0.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-500 kcal/100 grams of powder and theinstant beverage powder comprises at least 85% w/w BLG relative to totalprotein, preferably at least 90% w/w BLG relative to total protein, andthe pH of the powder is in the range of 6.1-8.5.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-500 kcal/100 grams of powder and theinstant beverage powder comprises at least 85% w/w BLG relative to totalprotein, preferably at least 90% w/w BLG relative to total protein, andthe pH of the powder is in the range of 2-8 and the instant beveragepowder further comprises vitamins, minerals and trace elements

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-500 kcal/100 grams of powder and theinstant beverage powder comprises at least 85% w/w BLG relative to totalprotein, preferably at least 90% w/w BLG relative to total protein, andthe pH of the powder is in the range of 2.0-4.9 and the instant beveragepowder further comprises vitamins, minerals and trace elements.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-500 kcal/100 grams of powder and theinstant beverage powder comprises at least 85% w/w BLG relative to totalprotein, preferably at least 90% w/w BLG relative to total protein andthe pH of the powder is in the range of 5.0-6.0 and the instant beveragepowder further comprises vitamins, minerals and trace elements.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-500 kcal/100 grams of powder and theinstant beverage powder comprises at least 85% w/w BLG relative to totalprotein, preferably at least 90% w/w BLG relative to total protein andthe pH of the powder is in the range of 6.1-8.5 and the instant beveragepowder further comprises vitamins, minerals and trace elements.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-500 kcal/100 grams of powder and theinstant beverage powder comprises at least 85% w/w BLG relative to totalprotein, preferably at least 90% w/w BLG relative to total protein, thepH of the powder is in the range of 2-8, the instant beverage powderfurther comprises vitamins, minerals and trace elements and wherein thesum of the amounts of Na, K, Mg, and Ca of the instant beverage powderis at most 10 mmol/g protein.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-500 kcal/100 grams of powder and theinstant beverage powder comprises at least 85% w/w BLG relative to totalprotein, preferably at least 90% w/w BLG relative to total protein, thepH of the powder is in the range of 2.0-4.9, the instant beverage powderfurther comprises vitamins, minerals and trace elements and wherein thesum of the amounts of Na, K, Mg, and Ca of the instant beverage powderis at most 10 mmol/g protein.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-500 kcal/100 grams of powder and theinstant beverage powder comprises at least 85% w/w BLG relative to totalprotein, preferably at least 90% w/w BLG relative to total protein, thepH of the powder is in the range of 5.0-6.0, the instant beverage powderfurther comprises vitamins, minerals and trace elements and wherein thesum of the amounts of Na, K, Mg, and Ca of the instant beverage powderis at most 10 mmol/g protein.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-500 kcal/100 grams of powder and theinstant beverage powder comprises at least 85% w/w BLG relative to totalprotein, preferably at least 90% w/w BLG relative to total protein, thepH of the powder is in the range of 6.1-8.5, the instant beverage powderfurther comprises vitamins, minerals and trace elements and wherein thesum of the amounts of Na, K, Mg, and Ca of the instant beverage powderis at most 10 mmol/g protein.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 400-500 kcal/100 grams of powder and theenergy distribution is in the range of 7-25 E % protein, 30-50 E %carbohydrate and 30-55 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 400-500 kcal/100 grams of powder and theenergy distribution is in the range of 7-25 E % protein, 30-50 E %carbohydrate and 30-55 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein and the pH of the powder isin the range of 2-8.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 400-500 kcal/100 grams of powder and theenergy distribution is in the range of 7-25 E % protein, 30-50 E %carbohydrate and 30-55 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein and the pH of the powder isin the range of 2.0-4.9.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 400-500 kcal/100 grams of powder and theenergy distribution is in the range of 7-25 E % protein, 30-50 E %carbohydrate and 30-55 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein and the pH of the powder isin the range of 5.0-6.0.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 400-500 kcal/100 grams of powder and theenergy distribution is in the range of 7-25 E % protein, 30-50 E %carbohydrate and 30-55 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein, and the pH of the powder isin the range of 6.1-8.5.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 400-500 kcal/100 grams of powder and theenergy distribution is in the range of 7-25 E % protein, 30-50 E %carbohydrate and 30-55 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein, and the pH of the powder isin the range of 2-8 and the instant beverage powder further comprisesvitamins, minerals and trace elements

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 400-500 kcal/100 grams of powder and theenergy distribution is in the range of 7-25 E % protein, 30-50 E %carbohydrate and 30-55 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein, and the pH of the powder isin the range of 2.0-4.9 and the instant beverage powder furthercomprises vitamins, minerals and trace elements.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 400-500 kcal/100 grams of powder and theenergy distribution is in the range of 7-25 E % protein, 30-50 E %carbohydrate and 30-55 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein and the pH of the powder isin the range of 5.0-6.0 and the instant beverage powder furthercomprises vitamins, minerals and trace elements.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 400-500 kcal/100 grams of powder and theenergy distribution is in the range of 7-25 E % protein, 30-50 E %carbohydrate and 30-55 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein and the pH of the powder isin the range of 6.1-8.5 and the instant beverage powder furthercomprises vitamins, minerals and trace elements.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 400-500 kcal/100 grams of powder and theenergy distribution is in the range of 7-25 E % protein, 30-50 E %carbohydrate and 30-55 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein, the pH of the powder is inthe range of 2-8, the instant beverage powder further comprisesvitamins, minerals and trace elements and wherein the sum of the amountsof Na, K, Mg, and Ca of the instant beverage powder is at most 10 mmol/gprotein.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 400-500 kcal/100 grams of powder and theenergy distribution is in the range of 7-25 E % protein, 30-50 E %carbohydrate and 30-55 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein, the pH of the powder is inthe range of 2.0-4.9, the instant beverage powder further comprisesvitamins, minerals and trace elements and wherein the sum of the amountsof Na, K, Mg, and Ca of the instant beverage powder is at most 10 mmol/gprotein.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 400-500 kcal/100 grams of powder and theenergy distribution is in the range of 7-25 E % protein, 30-50 E %carbohydrate and 30-55 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein, the pH of the powder is inthe range of 5.0-6.0, the instant beverage powder further comprisesvitamins, minerals and trace elements and wherein the sum of the amountsof Na, K, Mg, and Ca of the instant beverage powder is at most 10 mmol/gprotein.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 400-500 kcal/100 grams of powder and theenergy distribution is in the range of 7-25 E % protein, 30-50 E %carbohydrate and 30-55 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein, the pH of the powder is inthe range of 6.1-8.5, the instant beverage powder further comprisesvitamins, minerals and trace elements and wherein the sum of the amountsof Na, K, Mg, and Ca of the instant beverage powder is at most 10 mmol/gprotein.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-400 kcal/100 grams of powder and theenergy distribution is in the range of 10-30 E % protein, 70-90 E %carbohydrate and 0-5 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-400 kcal/100 grams of powder and theenergy distribution is in the range of 10-30 E % protein, 70-90 E %carbohydrate and 0-5 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% % w/w BLG relative to total protein and the pH of the powderis in the range of 2-8.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-400 kcal/100 grams of powder and theenergy distribution is in the range of 10-30 E % protein, 70-90 E %carbohydrate and 0-5 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein and the pH of the powder isin the range of 2.0-4.9.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-400 kcal/100 grams of powder and theenergy distribution is in the range of 10-30 E % protein, 70-90 E %carbohydrate and 0-5 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein and the pH of the powder isin the range of 5.0-6.0.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-400 kcal/100 grams of powder and theenergy distribution is in the range of 10-30 E % protein, 70-90 E %carbohydrate and 0-5 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein and the pH of the powder isin the range of 6.1-8.5.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-400 kcal/100 grams of powder and theenergy distribution is in the range of 10-30 E % protein, 70-90 E %carbohydrate and 0-5 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein and the pH of the powder isin the range of 2-8 and the instant beverage powder further comprisesvitamins, minerals and trace elements.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-400 kcal/100 grams of powder and theenergy distribution is in the range of 10-30 E % protein, 70-90 E %carbohydrate and 0-5 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein and the pH of the powder isin the range of 2.0-4.9 and the instant beverage powder furthercomprises vitamins, minerals and trace elements.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-400 kcal/100 grams of powder and theenergy distribution is in the range of 10-30 E % protein, 70-90 E %carbohydrate and 0-5 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein and the pH of the powder isin the range of 5.0-6.0 and the instant beverage powder furthercomprises vitamins, minerals and trace elements.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-400 kcal/100 grams of powder and theenergy distribution is in the range of 10-30 E % protein, 70-90 E %carbohydrate and 0-5 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein and the pH of the powder isin the range of 6.1-8.5 and the instant beverage powder furthercomprises vitamins, minerals and trace elements.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-400 kcal/100 grams of powder and theenergy distribution is in the range of 10-30 E % protein, 70-90 E %carbohydrate and 0-5 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein, the pH of the powder is inthe range of 2-8, the instant beverage powder further comprisesvitamins, minerals and trace elements and wherein the sum of the amountsof Na, K, Mg, and Ca of the instant beverage powder is at most 10 mmol/gprotein.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-400 kcal/100 grams of powder and theenergy distribution is in the range of 10-30 E % protein, 70-90 E %carbohydrate and 0-5 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein, the pH of the powder is inthe range of 2.0-4.9, the instant beverage powder further comprisesvitamins, minerals and trace elements and wherein the sum of the amountsof Na, K, Mg, and Ca of the instant beverage powder is at most 10 mmol/gprotein.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-400 kcal/100 grams of powder and theenergy distribution is in the range of 10-30 E % protein, 70-90 E %carbohydrate and 0-5 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein, the pH of the powder is inthe range of 5.0-6.0, the instant beverage powder further comprisesvitamins, minerals and trace elements and wherein the sum of the amountsof Na, K, Mg, and Ca of the instant beverage powder is at most 10 mmol/gprotein.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-400 kcal/100 grams of powder and theenergy distribution is in the range of 10-30 E % protein, 70-90 E %carbohydrate and 0-5 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein, the pH of the powder is inthe range of 6.1-8.5, the instant beverage powder further comprisesvitamins, minerals and trace elements and wherein the sum of the amountsof Na, K, Mg, and Ca of the instant beverage powder is at most 10 mmol/gprotein.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-350 kcal/100 grams of powder and theenergy distribution is in the range of 80-98 E % protein, 0-20 E %carbohydrate and 0-5 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-350 kcal/100 grams of powder and theenergy distribution is in the range of 80-98 E % protein, 0-20 E %carbohydrate and 0-5 E % lipid wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein and the pH of the powder isin the range of 2-8.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-350 kcal/100 grams of powder and theenergy distribution is in the range of 80-98 E % protein, 0-20 E %carbohydrate and 0-5 E % lipid, wherein the instant beverage powdercomprises at least 95% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein and the pH of the powder isin the range of 2.0-4.9.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-350 kcal/100 grams of powder and theenergy distribution is in the range of 80-98 E % protein, 0-20 E %carbohydrate and 0-5 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein and the pH of the powder isin the range of 5.0-6.0.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-350 kcal/100 grams of powder and theenergy distribution is in the range of 80-98 E % protein, 0-20 E %carbohydrate and 0-5 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein and the pH of the powder isin the range of 6.1-8.5.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-350 kcal/100 grams of powder and theenergy distribution is in the range of 80-98 E % protein, 0-20 E %carbohydrate and 0-5 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein and the pH of the powder isin the range of 2-8 and the instant beverage powder further comprisesvitamins, minerals and trace elements.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-350 kcal/100 grams of powder and theenergy distribution is in the range of 80-98 E % protein, 0-20 E %carbohydrate and 0-5 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein and the pH of the powder isin the range of 2.0-4.9 and the instant beverage powder furthercomprises vitamins, minerals and trace elements.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-350 kcal/100 grams of powder and theenergy distribution is in the range of 80-98 E % protein, 0-20 E %carbohydrate and 0-5 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein and the pH of the powder isin the range of 5.0-6.0 and the instant beverage powder furthercomprises vitamins, minerals and trace elements.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-350 kcal/100 grams of powder and theenergy distribution is in the range of 80-98 E % protein, 0-20 E %carbohydrate and 0-5 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein and the pH of the powder isin the range of 6.1-8.5 and the instant beverage powder furthercomprises vitamins, minerals and trace elements.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-350 kcal/100 grams of powder and theenergy distribution is in the range of 80-98 E % protein, 0-20 E %carbohydrate and 0-5 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein, the pH of the powder is inthe range of 2-8, the instant beverage powder further comprisesvitamins, minerals and trace elements and wherein the sum of the amountsof Na, K, Mg, and Ca of the instant beverage powder is at most 10 mmol/gprotein.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-350 kcal/100 grams of powder and theenergy distribution is in the range of 80-98 E % protein, 0-20 E %carbohydrate and 0-5 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein, the pH of the powder is inthe range of 2.0-4.9, the instant beverage powder further comprisesvitamins, minerals and trace elements and wherein the sum of the amountsof Na, K, Mg, and Ca of the instant beverage powder is at most 10 mmol/gprotein.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-350 kcal/100 grams of powder and theenergy distribution is in the range of 80-98 E % protein, 0-20 E %carbohydrate and 0-5 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein, the pH of the powder is inthe range of 5.0-6.0, the instant beverage powder further comprisesvitamins, minerals and trace elements and wherein the sum of the amountsof Na, K, Mg, and Ca of the instant beverage powder is at most 10 mmol/gprotein.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-350 kcal/100 grams of powder and theenergy distribution is in the range of 80-98 E % protein, 0-20 E %carbohydrate and 0-5 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein, the pH of the powder is inthe range of 6.1-8.5, the instant beverage powder further comprisesvitamins, minerals and trace elements and wherein the sum of the amountsof Na, K, Mg, and Ca of the instant beverage powder is at most 10 mmol/gprotein.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-420 kcal/100 grams of powder and theenergy distribution is in the range of 30-80 E % protein, 3-20 E %carbohydrate and 15-20 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-420 kcal/100 grams of powder and theenergy distribution is in the range of 30-80 E % protein, 3-20 E %carbohydrate and 15-20 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein and the pH of the powder isin the range of 2-8.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-420 kcal/100 grams of powder and theenergy distribution is in the range of 30-80 E % protein, 3-20 E %carbohydrate and 15-20 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein and the pH of the powder isin the range of 2.0-4.9.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-420 kcal/100 grams of powder and theenergy distribution is in the range of 30-80 E % protein, 3-20 E %carbohydrate and 15-20 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein and the pH of the powder isin the range of 5.0-6.0.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-420 kcal/100 grams of powder and theenergy distribution is in the range of 30-80 E % protein, 3-20 E %carbohydrate and 15-20 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein and the pH of the powder isin the range of 6.1-8.5.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-420 kcal/100 grams of powder and theenergy distribution is in the range of 30-80 E % protein, 3-20 E %carbohydrate and 15-20 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein and the pH of the powder isin the range of 2-8 and the instant beverage powder further comprisesvitamins, minerals and trace elements.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-420 kcal/100 grams of powder and theenergy distribution is in the range of 30-80 E % protein, 3-20 E %carbohydrate and 15-20 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein and the pH of the powder isin the range of 2.0-4.9 and the instant beverage powder furthercomprises vitamins, minerals and trace elements.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-420 kcal/100 grams of powder and theenergy distribution is in the range of 30-80 E % protein, 3-20 E %carbohydrate and 15-20 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein and the pH of the powder isin the range of 5.0-6.0 and the instant beverage powder furthercomprises vitamins, minerals and trace elements.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-420 kcal/100 grams of powder and theenergy distribution is in the range of 30-80 E % protein, 3-20 E %carbohydrate and 15-20 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% % w/w BLG relative to total protein and the pH of the powderis in the range of 6.1-8.5 and the instant beverage powder furthercomprises vitamins, minerals and trace elements.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-420 kcal/100 grams of powder and theenergy distribution is in the range of 30-80 E % protein, 3-20 E %carbohydrate and 15-20 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein, the pH of the powder is inthe range of 2-8, the instant beverage powder further comprisesvitamins, minerals and trace elements and wherein the sum of the amountsof Na, K, Mg, and Ca of the instant beverage powder is at most 10 mmol/gprotein.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-420 kcal/100 grams of powder and theenergy distribution is in the range of 30-80 E % protein, 3-20 E %carbohydrate and 15-20 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein, the pH of the powder is inthe range of 2.0-4.9, the instant beverage powder further comprisesvitamins, minerals and trace elements and wherein the sum of the amountsof Na, K, Mg, and Ca of the instant beverage powder is at most 10 mmol/gprotein.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-420 kcal/100 grams of powder and theenergy distribution is in the range of 30-80 E % protein, 3-20 E %carbohydrate and 15-20 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein, the pH of the powder is inthe range of 5.0-6.0, the instant beverage powder further comprisesvitamins, minerals and trace elements and wherein the sum of the amountsof Na, K, Mg, and Ca of the instant beverage powder is at most 10 mmol/gprotein.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 200-420 kcal/100 grams of powder and theenergy distribution is in the range of 30-80 E % protein, 3-20 E %carbohydrate and 15-20 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein, the pH of the powder is inthe range of 6.1-8.5, the instant beverage powder further comprisesvitamins, minerals and trace elements and wherein the sum of the amountsof Na, K, Mg, and Ca of the instant beverage powder is at most 10 mmol/gprotein.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 150-250 kcal/100 grams of powder and theenergy distribution is in the range of 10-30 E % protein, 40-60 E %carbohydrate and 25-45 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 150-250 kcal/100 grams of powder and theenergy distribution is in the range of 10-30 E % protein, 40-60 E %carbohydrate and 25-45 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein and the pH of the powder isin the range of 2-8.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 150-250 kcal/100 grams of powder and theenergy distribution is in the range of 10-30 E % protein, 40-60 E %carbohydrate and 25-45 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein and the pH of the powder isin the range of 2.0-4.9.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 150-250 kcal/100 grams of powder and theenergy distribution is in the range of 10-30 E % protein, 40-60 E %carbohydrate and 25-45 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein and the pH of the powder isin the range of 5.0-6.0.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 150-250 kcal/100 grams of powder and theenergy distribution is in the range of 10-30 E % protein, 40-60 E %carbohydrate and 25-45 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein and the pH of the powder isin the range of 6.1-8.5.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 150-250 kcal/100 grams of powder and theenergy distribution is in the range of 10-30 E % protein, 40-60 E %carbohydrate and 25-45 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein and the pH of the powder isin the range of 2-8 and the instant beverage powder further comprisesvitamins, minerals and trace elements.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 150-250 kcal/100 grams of powder and theenergy distribution is in the range of 10-30 E % protein, 40-60 E %carbohydrate and 25-45 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein and the pH of the powder isin the range of 2.0-4.9 and the instant beverage powder furthercomprises vitamins, minerals and trace elements.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 150-250 kcal/100 grams of powder and theenergy distribution is in the range of 10-30 E % protein, 40-60 E %carbohydrate and 25-45 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein and the pH of the powder isin the range of 5.0-6.0 and the instant beverage powder furthercomprises vitamins, minerals and trace elements.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 150-250 kcal/100 grams of powder and theenergy distribution is in the range of 10-30 E % protein, 40-60 E %carbohydrate and 25-45 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% % w/w BLG relative to total protein and the pH of the powderis in the range of 6.1-8.5 and the instant beverage powder furthercomprises vitamins, minerals and trace elements.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 150-250 kcal/100 grams of powder and theenergy distribution is in the range of 10-30 E % protein, 40-60 E %carbohydrate and 25-45 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein, the pH of the powder is inthe range of 2-8, the instant beverage powder further comprisesvitamins, minerals and trace elements and wherein the sum of the amountsof Na, K, Mg, and Ca of the instant beverage powder is at most 10 mmol/gprotein.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 150-250 kcal/100 grams of powder and theenergy distribution is in the range of 10-30 E % protein, 40-60 E %carbohydrate and 25-45 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein, the pH of the powder is inthe range of 2.0-4.9, the instant beverage powder further comprisesvitamins, minerals and trace elements and wherein the sum of the amountsof Na, K, Mg, and Ca of the instant beverage powder is at most 10 mmol/gprotein.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 150-250 kcal/100 grams of powder and theenergy distribution is in the range of 10-30 E % protein, 40-60 E %carbohydrate and 25-45 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein, the pH of the powder is inthe range of 5.0-6.0, the instant beverage powder further comprisesvitamins, minerals and trace elements and wherein the sum of the amountsof Na, K, Mg, and Ca of the instant beverage powder is at most 10 mmol/gprotein.

In one embodiment of the invention, the instant beverage powder has anenergy content in the range of 150-250 kcal/100 grams of powder and theenergy distribution is in the range of 10-30 E % protein, 40-60 E %carbohydrate and 25-45 E % lipid, wherein the instant beverage powdercomprises at least 85% w/w BLG relative to total protein, preferably atleast 90% w/w BLG relative to total protein, the pH of the powder is inthe range of 6.1-8.5, the instant beverage powder further comprisesvitamins, minerals and trace elements and wherein the sum of the amountsof Na, K, Mg, and Ca of the instant beverage powder is at most 10 mmol/gprotein.

In some preferred embodiments of the invention the food product is a dryfood product, e.g. a bar or an instant beverage powder, comprisingcarbohydrate and protein, said dry food product comprising at least 1%w/w BLG, preferably at least 5%, wherein:

i) the crystallinity of BLG is at least 20%, preferably at least 40%,and/orii) at least 90% w/w of the total amount of protein is comprised by BLG.

In some particularly preferred embodiments of the invention the foodproduct is a low phosphorus food product comprising at most 40 mgphosphorus per 100 g protein.

Non-limiting examples of the food product are e.g. a dairy product, acandy, a beverage, an instant beverage, a protein bar, an enteralnutritional composition, a bakery product.

In other preferred embodiments of the invention the food product is aninstant beverage powder comprising, or even essentially consisting of:

-   -   an edible BLG composition as defined herein in the form of a        powder to provide at total amount of BLG of at least 1% w/w,        preferably at least 5% w/w, said edible BLG composition having a        crystallinity of BLG of at least 20%,    -   a sweetener in the form of a powder, e.g. a sugar sweetener        and/or a non-sugar sweetener,    -   optionally, a flavouring agent    -   at least one food acid in the form of a powder, e.g. citric acid        or other suitable food acids, and    -   at most 80 mg phosphorus/100 g protein, and        PS wherein a 10% solution of the instant beverage powder in        demineralised water has a pH in the range of 2.5-4.0.

In some preferred embodiments of the invention, the edible BLGcomposition comprises:

-   -   At most 6% w/w water    -   At least 80% total protein relative to total solids    -   At least 95% BLG relative to total protein, and        said edible BLG composition:    -   Is a dry powder, and    -   Has a bulk density of at least 0.50 g/mL, and preferably at        least 0.60 g/mL.

In other preferred embodiments of the invention, the edible BLGcomposition comprises:

-   -   At most 6% w/w water    -   At least 80% total protein relative to total solids    -   At least 95% BLG relative to total protein, and        said edible BLG composition:    -   Is a dry powder,    -   Has a bulk density of at least 0.50 g/mL, and preferably at        least 0.60 g/mL, and    -   Has a crystallinity of BLG of at least 20% and preferably at        least 40%.

In further preferred embodiments of the invention, the edible BLGcomposition comprises:

-   -   At most 6% w/w water    -   At least 80% total protein relative to total solids,    -   At least 95% BLG relative to total protein,    -   at most 80 mg phosphorus per 100 g protein.        said edible BLG composition:    -   Is a dry powder.

In yet preferred embodiments of the invention, the edible BLGcomposition comprises:

-   -   At most 6% w/w water    -   At least 90% total protein relative to total solids,        At least 97% BLG relative to total protein,    -   at most 50 mg phosphorus per 100 g protein.        said edible BLG composition:    -   Is a dry powder.

In other preferred embodiments of the invention, the edible BLGcomposition comprises:

-   -   At most 6% w/w water    -   At least 80% total protein relative to total solids, and        preferably at least 90% total protein relative to total solids,    -   30-70% BLG relative to total protein,    -   8-25% w/w ALA relative to total protein,        said edible BLG composition:    -   Is a dry powder, and    -   Has a crystallinity of BLG of at least 20% and preferably at        least 40%.

In one aspect of the invention, a liquid food product is prepared fromthe instant beverage powder. By use of the instant beverage powderaccording to the invention, it is possible to obtain a liquid foodproduct within a very short time.

Thus, in one aspect of the invention pertains to a method for preparinga liquid food product according to the invention, said method comprising

-   -   i. Adding an instant beverage powder according to the invention,    -   ii. Optionally adding at least one further ingredient, and    -   iii. Mixing the powder and liquid obtained to form a uniform        mixture.

In one embodiment the liquid is selected from the group consisting ofwater, milk products, fruit juice, vegetable juice, beverages andcombinations thereof. In one embodiment, the further ingredient isselected from fruits or vegetables.

When mixing the powder and liquid, the mixing can be performed byshaking. After shaking, the liquid food product, the instant beveragepowder may be allowed to stand for ½-2 minutes in order to fullydissolve. One advantage of the instant beverage powder is that theinstant beverage powder easily dissolves, forms a uniform solution andremains a uniform solution, i.e. substantially no segregation occurs.

A problem usually associated with instant beverage powders is that whenpreparing a liquid food product from the powder, foam is developed. Theinstant beverage powder of the invention has a tendency not to foam whenpreparing a liquid food product from the powder.

A liquid food product comprising a liquid and the powder according tothe invention may be prepared by mixing the instant beverage powder ofthe invention with the liquid. In one embodiment of the invention, theinstant beverage powder may comprise at most 40 gram of said powder per100 grams of said liquid, such as at most 30 gram of said powder per 100grams of said liquid. In a preferred embodiment of the invention, theliquid food product comprises 1-30 gram of said powder per 100 grams ofsaid liquid, more preferably 1-20 gram of said powder, even morepreferably 1-10 gram of said powder, or preferably 1-5 gram of saidpowder or even more preferably 2.5-5 gram of said powder.

In one embodiment of the invention the liquid food product comprises5-25 gram of said powder per 100 gram of the liquid, preferably 5-25gram of said powder per 100 gram of the liquid, more preferably 10-15gram of said powder, even more preferably 11-14 gram of said powder ormore preferably 11-12 gram of said powder.

In one embodiment of the invention the food product has energy contentin the range of 30-300 kcal/100 grams of food product, preferably in therange of 30-100 kcal/100 grams of food product, more preferably in therange of 40-90 kcal/100 grams of food product, or even more preferablyin the range of 40-70 kcal/100 grams of food product.

Alternatively, the food product has energy content in the range of100-300 kcal/100 grams of food product, preferably in the range of100-250 kcal/100 grams of food product, or more preferably in the rangeof 125-225 kcal/100 grams of food product.

The liquid food product may comprise a liquid selected from the groupconsisting of water, milk products, fruit juice, vegetable juice,beverages and combinations thereof.

The appearance of liquid food products e.g. a beverage prepared from theinstant beverage powder, is of great importance to the consumer.Transparency is a parameter that the consumer uses to evaluate theproduct. One way of determining the transparency of the liquid foodproduct is by measuring the turbidity of the product as described inexample 1.7.

In some embodiments of the beverage prepared from the instant beveragepowder, it is beneficial that the beverage is transparent. This may forexample be advantageous when the beverage is used as a sport beverage orin “protein water”, in which case it is beneficial that the beverageresemble water in appearance.

In a preferred embodiment of the present invention, the beverageprepared from the instant beverage powder has a turbidity of at most 200NTU, and such a beverage is transparent and/or translucent.

In some preferred embodiment of the present invention, the beverageprepared from the instant beverage powder have a turbidity of at most150 NTU, or preferably a turbidity of at most 100 NTU, or preferably aturbidity of at most 80 NTU, or preferably a turbidity of at most 60 NTUor more preferably a turbidity of at most 40 NTU, or preferably aturbidity of at most 30 NTU, preferably a turbidity of at most 20 NTU,more preferably a turbidity of at most 10 NTU, and more preferably aturbidity of at most 5 NTU, even more preferably, it has a turbidity ofat most 2 NTU.

In a preferred embodiment of the present invention the beverage preparedfrom the instant beverage powder have a turbidity of more than 200 NTU,such a beverage is opaque.

In some embodiments of the beverage prepared from the instant beveragepowder, it is beneficial that the beverage is opaque. This is forexample advantageous when the beverage should resemble milk and have amilky appearance. The appearance of nutritionally complete nutritionalsupplements is typically opaque.

In some preferred embodiments of the invention, the beverage preparedfrom the instant beverage powder have a turbidity of more than 250 NTU.Preferably the beverage have a turbidity of more than 300 NTU, morepreferably, it has a turbidity of more than 500 NTU, more preferably ithas a turbidity of more than 1000, preferably a turbidity of more than1500 NTU, even more preferably it has a turbidity of more than 2000 NTU.

The colour of the product is of great importance to the consumer.Instant beverage powder products comprising whey proteins have aslightly yellow colour. However, when using instant beverage powdercomprising BLG having a crystallinity of at least 20% or instantbeverage powder, where at least 90% of the protein is comprised by BLG,the colour of the product is substantially less yellow and the productappears more white. Thus, addition of colour to the instant beveragepowder in order to mask the yellow colour is not necessary.

One way of measuring the colour of a product is by using the CIELABcolour space, which expresses colour as three numerical values; L* forlightness and a* and b* for the green-red and blue-yellow colourcomponents. Example 1.9 describes how to measure the L*, a* and b*values for the liquid food product.

In one embodiment of the invention the protein fraction of the liquidfood product has a colour value delta b* in the range of −0.10 to +0.51at the CIELAB color scale, wherein deltab*=b_(sample standardized to 6.0 w/w % protein)*−b_(demin. water)*,measured at room temperature.

Another aspect of the invention pertains to a method for preparing aninstant beverage powder comprising BLG and at least one optionalingredient, said method comprising blending a dry BLG isolate, with atleast one additional ingredient selected from the group consisting ofvitamins, flavouring agent, colouring agent, minerals, sweeteners,antioxidants, food acid, lipids, carbohydrate, prebiotics, probiotics,anti-foaming agents and non-whey protein to obtain an instant beveragepowder.

In one embodiment of the invention, the BLG of the BLG source is coatedwith an organic acid. If the BLG source e.g. is a powder, this meansthat the powder is coated with the organic acid. When preparing instantbeverage powders comprising protein, lecithin is commonly used forimproving the solubility of the protein. However, lecithin is a sourceof phosphorus. It is therefore desirable to find another way ofimproving the solubility of instant beverage powders comprising protein.

The inventors have found that by coating the BLG crystals or powderparticles of the BLG source with one or more organic acids, thesolubility of the instant beverage powder improves. The organic acid orsalt of organic acid can be selected from the group consisting ofpyruvate, aconitate, citrate, iso-citrate, ketoglutarate, succinyl-CoA,succinate, fumarate, malate, oxaloacetate, tartrate, acetate, tannicacid, benzoic acid, maleic acid and lactate. In a preferred embodimentof the invention, the BLG crystals are coated with organic acid or saltof organic acid selected from the group consisting of pyruvate, citrate,iso-citrate, ketoglutarate, succinate, fumarate, malate, oxaloacetate,tartrate, acetate, maleic acid and lactate and salts thereof

In a preferred embodiment of the invention, the BLG crystals of the BLGsource are coated with citrate, e.g. a citrate selected from the groupconsisting of trisodium citrate, potassium citrate and calcium citrate.

In preferred embodiment of the invention, the BLG of the BLG source iscoated with organic acid by use of spray-drying or fluid bed. It isparticularly preferred that dried BLG crystals are are coated with theorganic acid or salts thereof using e.g. spray-drying or fluid bed.

The BLG source can be obtained from a whey protein feed, from which theBLG is isolated as crystals. One method of preparing the BLG isolate isdescribed in international patent application No. PCT/EP2017/084553,which is hereby incorporated by reference. The BLG isolate can beprepared as described on page 6, line 23-32 of PCT/EP2017/084553 asfiled, where the edible composition comprising BLG in crystallisedand/or isolated form corresponds to the BLG isolate of the presentinvention. In a preferred embodiment, the BLG isolate is prepared by themethod described on page 39, line 15-34 of PCT/EP2017/084553 as filed.In another preferred embodiment the BLG isolate is prepared by themethod described on page 41, line 1-24 of PCT/EP2017/084553 as filed.

In some preferred embodiments of the invention the instant beveragepowder comprises, or even consists of, a BLG isolate powder comprisingdried BLG crystals, said BLG isolate powder is coated with an organicacid and/or a salt of an organic acid. The weight ratio between theweight of the BLG isolate and the total weight of the sum of organicacids and deprotonated organic acids is preferably 5-100, morepreferably 8-60, even more preferably 10-40, and most preferably 12-30.

An aspect of the invention pertains to a method of producing a BLGisolate powder coated with organic acid and/or a salt of an organicacid, the method comprises the following steps

-   -   Providing a BLG isolate powder to be coated, preferably        comprising, or even consisting of, dried BLG crystals,        preferably obtained by the BLG crystallization process described        herein, and    -   Applying organic acid and/or salt of an organic acid to the BLG        isolate powder to be coated, preferably in an amount sufficient        to coat the BLG isolate powder but avoiding that it is        dissolved,    -   Optionally evaporating residual moisture from the coated BLG        isolate powder.

The BLG isolate powder to be coated, preferably has both a high proteincontent and a high BLG purity. The BLG isolate powder to be coatedpreferably has a crystallinity of BLG of at least 20%, preferably atleast 40%, more preferably at least 60%, and even more preferably atleast 80%.

The organic acid and/or salt of an organic acid to the BLG isolatepowder to be coated in an amount sufficient to provide a weight ratiobetween the weight of the BLG isolate powder and the total weight of thesum of organic acids and deprotonated organic acids is preferably 5-100,more preferably 8-60, even more preferably 10-40, and most preferably12-30.

The organic acid and/or salt of a organic acid is preferably applied tothe BLG isolate powder in a fluid bed system by spraying organic acidand/or salt of an organic acid, preferably in dissolved form into thefluid bed to coat the BLG isolate powder. The temperature duringoperation is preferably in the range of 5-70 degrees C. more preferablyin the range of 50-65 degrees C. such as preferably approx. 60 degreesC.

After application of the organic acid and/or salt of an organic acid thecoated BLG isolate may be processed to evaporate additional moisture,preferably until the water content is at most 6% w/w and more preferablyat most 5% w/w.

The organic acids are preferably edible organic acids, i.e. so-calledfood acids.

In some preferred embodiments of the invention the BLG source used forpreparing the instant beverage powder has a solids content of at least20% w/w. Preferably, the BLG source has a solids content of at least 30%w/w, more preferably, the BLG source has a solids content of at least40% w/w, even more preferably, the BLG source has a solids content of atleast 50% w/w, such as e.g. at least 60% w/w.

In other preferred embodiments of the invention the BLG source used forpreparing the instant beverage powder has a solid content of in therange of 20-80% w/w. Preferably, the BLG source has a solid content inthe range of 30-70% w/w. More preferably, the BLG source has a solidcontent in the range of 40-65% w/w. Even more preferably, the BLG sourcehas a solid content in the range of 50-65% w/w, such as e.g. approx. 60%w/w.

The BLG source is preferably a BLG isolate powder or a liquid BLGisolate contain water and the solids of the BLG isolate powder in anamount in the range from 1-50% w/w. It is particularly preferred thatthe BLG source is a BLG isolate powder.

The beta-lactoglobulin (BLG) isolate powder, preferably prepared byspray-drying, has a pH in the range of i) 2-4.9, ii) 6.1-8.5, or iii)5.0-6.0 and comprises:

-   -   total protein in an amount of at least 30% w/w,    -   BLG in an amount of at least 85% w/w relative to total protein,        and    -   water in an amount of at most 10% w/w.

The BLG isolate powder preferably has one or more of the following:

-   -   a bulk density of at least 0.2 g/cm³,    -   an intrinsic tryptophan fluorescence emission ratio (I330/I350)        of at least 1.11,    -   a degree of protein denaturation of at most 10%,    -   a heat-stability at pH 3.9 of at most 200 NTU, and    -   at most 1000 colony-forming units/g.

The BLG isolate powder is preferably an edible composition. In somepreferred embodiments of the invention the BLG isolate powder is anedible BLG composition as defined herein.

In some preferred embodiments of the invention, the BLG isolate powderhas a pH in the range of 2-4.9. Such powders are particularly useful foracidic food products and particularly acidic beverages.

In other preferred embodiments of the invention, BLG isolate powder hasa pH in the range of 6.1-8.5.

In some preferred embodiments of the invention, the BLG isolate powdercomprises total protein in an amount of at least 40% w/w, preferably atleast 50% w/w, at least 60% w/w, more preferably at least 70% w/w, evenmore preferably at least 80% w/w.

Even higher protein contents may be required and in some preferredembodiments of the invention, the BLG isolate powder comprises totalprotein in an amount of at least 85% w/w, preferably at least 90% w/w,at least 92% w/w, more preferably at least 94% w/w, and even morepreferably at least 95% w/w.

Total protein is measured according to Example 1.5.

In some preferred embodiments of the invention, the BLG isolate powdercomprises BLG in an amount of at least 92% w/w relative to totalprotein, preferably at least 95% w/w, more preferably at least 97% w/w,even more preferably at least 98%, and most preferably BLG in an amountof at least 99.5% w/w relative to total protein.

In some preferred embodiments of the invention, the sum ofalpha-lactalbumin (ALA) and caseinomacropeptide (CMP) comprises at least40% w/w of the non-BLG protein of the powder, preferably at least 60%w/w, even more preferably at least 70% w/w, and most preferably at least90% w/w of the non-BLG protein of the powder.

In other preferred embodiments of the invention, each main non-BLG wheyprotein is present in a weight percentage relative to total proteinwhich is at most 25% of its weight percentage relative to total proteinin a standard whey protein concentrate from sweet whey, preferably atmost 20%, more preferably at most 15%, even more preferably at most 10%,most preferably at most 6%.

Even lower concentrations of the main non-BLG whey proteins may bedesirable. Thus, in additional preferred embodiments of the invention,each main non-BLG whey protein is present in a weight percentagerelative to total protein which is at most 4% of its weight percentagerelative to total protein in a standard whey protein concentrate fromsweet whey, preferably at most 3%, more preferably at most 2%, even morepreferably at most 1%.

The inventors have seen indications that reduction of lactoferrin and/orlactoperoxidase is particularly advantageous for obtaining acolour-neutral whey protein product.

Thus in some preferred embodiments of the invention, lactoferrin ispresent in a weight percentage relative to total protein which is atmost 25% of its weight percentage relative to total protein in astandard whey protein concentrate from sweet whey, preferably at most20%, more preferably at most 15%, even more preferably at most 10%, mostpreferably at most 6%. Even lower concentrations of lactoferrin may bedesirable. Thus, in additional preferred embodiments of the invention,lactoferrin is present in a weight percentage relative to total proteinwhich is at most 4% of its weight percentage relative to total proteinin a standard whey protein concentrate from sweet whey, preferably atmost 3%, more preferably at most 2%, even more preferably at most 1%.

Similarly, in some preferred embodiments of the invention,lactoperoxidase is present in a weight percentage relative to totalprotein which is at most 25% of its weight percentage relative to totalprotein in a standard whey protein concentrate from sweet whey,preferably at most 20%, more preferably at most 15%, even morepreferably at most 10%, most preferably at most 6%. Even lowerconcentrations of lactoperoxidase may be desirable. Thus, in additionalpreferred embodiments of the invention, lactoperoxidase is present in aweight percentage relative to total protein which is at most 4% of itsweight percentage relative to total protein in a standard whey proteinconcentrate from sweet whey, preferably at most 3%, more preferably atmost 2%, even more preferably at most 1%.

Lactoferrin and lactoperoxidase are quantified according to Example1.29.

In some preferred embodiments of the invention, the BLG isolate powderhas a water content in an amount of at most 10% w/w, preferably at most7% w/w, more preferably at most 6% w/w, even more preferably at most 4%w/w, and most preferred at most 2% w/w.

In some preferred embodiments of the invention the BLG isolate powdercomprises carbohydrate in an amount of at most 60% w/w, preferably atmost 50% w/w, more preferably at most 20% w/w, even more preferably atmost 10% w/w, even more preferably at most 1% w/w, and most preferablyat most 0.1%. The BLG isolate powder may for example containcarbohydrates, such as e.g. lactose, oligosaccharides and/or hydrolysisproducts of lactose (i.e. glucose and galactose), sucrose, and/ormaltodextrin.

In some preferred embodiments of the invention, the BLG isolate powdercomprises lipid in an amount of at most 10% w/w, preferably at most 5%w/w, more preferably at most 2% w/w, and even more preferably at most0.1% w/w.

The present inventors have found that it can be advantageous to controlthe mineral content to reach some of the desired properties of the BLGisolate powder.

In some preferred embodiments of the invention, the sum of the amountsof Na, K, Mg, and Ca of the BLG isolate powder is at most 10 mmol/gprotein. Preferably, the sum of the amounts of Na, K, Mg, and Ca of theBLG isolate powder is at most 6 mmol/g protein, more preferably at most4 mmol/g protein, even more preferably at most 2 mmol/g protein.

In other preferred embodiments of the invention, the the sum of theamounts of Na, K, Mg, and Ca of the BLG isolate powder is at most 1mmol/g protein. Preferably, the sum of the amounts of Na, K, Mg, and Caof the BLG isolate powder is at most 0.6 mmol/g protein, more preferablyat most 0.4 mmol/g protein, even more preferably at most 0.2 mmol/gprotein, and most preferably at most 0.1 mmol/g protein.

In other preferred embodiments of the invention, the sum of the amountsof Mg and Ca of the BLG isolate powder is at most 5 mmol/g protein.Preferably, the sum of the amounts of Mg and Ca of the BLG isolatepowder is at most 3 mmol/g protein, more preferably at most 1.0 mmol/gprotein, even more preferably at most 0.5 mmol/g protein.

In other preferred embodiments of the invention, the sum of the amountsof Mg and Ca of the BLG isolate powder is at most 0.3 mmol/g protein.Preferably, the sum of the amounts of Mg and Ca of the BLG isolatepowder is at most 0.2 mmol/g protein, more preferably at most 0.1 mmol/gprotein, even more preferably at most 0.03 mmol/g protein, and mostpreferably at most 0.01 mmol/g protein.

The inventors have found that it is possible to use low phosphorus/lowpotassium variants of the BLG isolate powder that are particularlyuseful to patients with kidney diseases. To make such a product, the BLGisolate powder has to have an equally low content of phosphorus andpotassium.

Thus, in some preferred embodiments of the invention, the BLG isolatepowder has a total content of phosphorus of at most 100 mg phosphorusper 100 g protein. Preferably, the BLG isolate powder has a totalcontent of at most 80 mg phosphorus per 100 g protein. More preferably,the BLG isolate powder has a total content of at most 50 mg phosphorusper 100 g protein. Even more preferably, the BLG isolate powder has atotal content of phosphorus of at most 20 mg phosphorus per 100 gprotein. The BLG isolate powder has a total content of phosphorus of atmost 5 mg phosphorus per 100 g protein.

In some preferred embodiments of the invention, the BLG isolate powdercomprises at most 600 mg potassium per 100 g protein. More preferably,the BLG isolate powder comprise at most 500 mg potassium per 100 gprotein. More preferably, the BLG isolate powder comprises at most 400mg potassium per 100 g protein. More preferably, the BLG isolate powdercomprises at most 300 mg potassium per 100 g protein. Even morepreferably, the BLG isolate powder at most 200 mg potassium per 100 gprotein. Even more preferably, the BLG isolate powder comprises at most100 mg potassium per 100 g protein. Even more preferably, the BLGisolate powder comprises at most 50 mg potassium per 100 g protein andeven more preferably, the BLG isolate powder comprises at most 10 mgpotassium per 100 g protein.

The content of phosphorus relates to the total amount of elementalphosphorus of the composition in question and is determined according toExample 1.19. Similarly, the content of potassium relates to the totalamount of elemental potassium of the composition in question and isdetermined according to Example 1.19.

In some preferred embodiments of the invention, the BLG isolate powdercomprises at most 100 mg phosphorus/100 g protein and at most 700 mgpotassium/100 g protein, preferably at most 80 mg phosphorus/100 gprotein and at most 600 mg potassium/100 g protein, more preferably atmost 60 mg phosphorus/100 g protein and at most 500 mg potassium/100 gprotein, more preferably at most 50 mg phosphorus/100 g protein and atmost 400 mg potassium/100 g protein, or more preferably at most 20 mgphosphorus/100 g protein and at most 200 mg potassium/100 g protein, oreven more preferably at most 10 mg phosphorus/100 g protein and at most50 mg potassium/100 g protein. In some preferred embodiments of theinvention the BLG isolate powder comprises at most 100 mg phosphor/100 gprotein and at most 340 mg potassium/100 g protein.

The low phosphorus and/or low potassium compositions according to thepresent invention may be used as a food ingredient for the production ofa food product for patients groups that have a reduced kidney function.

The present inventors have found that for some applications, e.g. acidicfood products and particularly acidic beverages, it is particularlyadvantageous to have an acidic BLG isolate powder having a pH of at most4.9 and even more preferably at most 4.3. This is especially true forhigh protein, transparent acidic beverages.

In the context of the present invention, a transparent liquid has aturbidity of at most 200 NTU measured according to Example 1.7.

Thus, in some preferred embodiments of the invention, the BLG isolatepowder has a pH in the range of 2-4.9. Preferably, the BLG isolatepowder has a pH in the range of 2.5-4.7, more preferably 2.8-4.3, evenmore preferably 3.2-4.0, and most preferably 3.4-3.9. Alternatively, butalso preferred, the BLG isolate powder may have a pH in the range of3.6-4.3.

The present inventors have found that for some applications, e.g.pH-neutral food products and particularly pH-neutral beverages, it isparticularly advantageous to have a pH-neutral BLG isolate powder. Thisis especially true for high protein, transparent or opaque pH-neutralbeverages.

Thus, in some preferred embodiments of the invention, BLG isolate powderhas a pH in the range of 6.1-8.5. Preferably, the powder has a pH in therange of 6.1-8.5, more preferably 6.2-8.0, even more preferably 6.3-7.7,and most preferably 6.5-7.5.

In other preferred embodiments of the invention, BLG isolate powder hasa pH in the range of 5.0-6.0. Preferably, the powder has a pH in therange of 5.1-5.9, more preferably 5.2-5.8, even more preferably 5.3-5.7,and most preferably 5.4-5.6.

Advantageously, the BLG isolate powder used in the present invention mayhave bulk density of at least 0.20 g/cm³, preferably at least 0.30g/cm³, more preferably at least 0.40 g/cm³, even more preferably atleast 0.45 g/cm³, even more preferably at least 0.50 g/cm³, and mostpreferably at least 0.6 g/cm³.

Low density powders such as freeze-dried BLG isolates are fluffy andeasily drawn into the air of the production site during use. This isproblematic as it increases the risk of cross-contamination of thefreeze-dried powder to other foods products and a dusty environment isknown to be a cause of hygiene issues. In extreme cases, a dustyenvironment also increases the risk of dust explosions.

The high density variants of the present invention are easier to handleand less prone to flow into the surrounding air.

An additional advantage of the high density variants of the presentinvention is that they take up less space during transportation andthereby increase weight of BLG isolate powder that can be transported inone volume unit.

Yet an advantage of the high density variants of the present inventionis that they are less prone to segregation when used in powder mixtureswith other powdered food ingredients, such as e.g. powdered sugar (bulkdensity of approx. 0.56 g/cm³), granulated sugar (bulk density ofapprox. 0.71 g/cm³), powdered citric acid (bulk density of approx. 0.77g/cm³).

The BLG isolate powder of the present invention may have bulk density inthe range of 0.2-1.0 g/cm³, preferably in the range of 0.30-0.9 g/cm³,more preferably in the range of 0.40-0.8 g/cm³, even more preferably inthe range of 0.45-0.75 g/cm³, even more preferably in the range of0.50-0.75 g/cm³, and most preferably in the range of 0.6-0.75 g/cm³.

The bulk density of a powder is measured according to Example 1.17.

The present inventors have found that it is advantageous to maintain thenative conformation of BLG and have seen indications that increasedunfolding of BLG gives rise to an increased level of drying mouthfeelwhen the BLG is used for acidic beverages.

The intrinsic tryptophan fluorescence emission ratio (I330/I350) is ameasure of degree of unfolding of BLG and the inventors have found thatat high intrinsic tryptophan fluorescence emission ratios, whichcorrelate with low or no unfolding of BLG, less drying mouthfeel wasobserved. The intrinsic tryptophan fluorescence emission ratio(I330/I350) is measured according to Example 1.1.

In some preferred embodiments of the invention, the BLG isolate powderhas an intrinsic tryptophan fluorescence emission ratio (I330/I350) ofat least 1.11.

In some preferred embodiments of the invention, the BLG isolate powderhas an intrinsic tryptophan fluorescence emission ratio (I330/I350) ofat least 1.12, preferably at least 1.13, more preferably at least 1.15,even more preferably at least 1.17, and most preferably at least 1.19.

If BLG isolate powder contains considerable amounts of non-proteinmatter it is preferred to isolate the protein fraction before measuringthe intrinsic tryptophan fluorescence emission ratio. Thus in somepreferred embodiments of the invention, the protein fraction of the BLGisolate powder has an intrinsic tryptophan fluorescence emission ratioof at least 1.11.

In some preferred embodiments of the invention, the protein fraction ofthe BLG isolate powder has an intrinsic tryptophan fluorescence emissionratio (I330/I350) of at least 1.12, preferably at least 1.13, morepreferably at least 1.15, even more preferably at least 1.17, and mostpreferably at least 1.19.

The protein fraction can e.g. be separated from the BLG isolate powderby dissolving the BLG isolate powder in demineralised water andsubjecting the solution to dialysis or ultrafiltration-baseddiafiltration using a filter that retains the protein. If the BLGisolate powder contains interfering levels of lipid such lipid can e.g.be removed by microfiltration. Steps of microfiltration andultrafiltration/diafiltration can be combined to remove both lipid andsmall molecules from the protein fraction.

It is often preferred that a substantial amount of the BLG of the BLGisolate powder is non-aggregated BLG. Preferably at least 50% of the BLGis non-aggregated BLG. More preferably at least at least 80% of the BLGis non-aggregated BLG. Even more preferred at least 90% of the BLG isnon-aggregated BLG. Most preferred, at least 95% of the BLG isnon-aggregated BLG. Even more preferred approx. 100% of the BLG of theBLG isolate powder is non-aggregated BLG.

In some preferred embodiments of the invention, the BLG isolate powderhas a degree of protein denaturation of at most 10%, preferably at most8%, more preferably at most 6%, even more preferably at most 3%, evenmore preferably at most 1%, and most preferably at most 0.2%.

However, it may also be preferred that the BLG isolate powder has asignificant level of protein denaturation, e.g. if an opaque beverage isdesired. Thus, in other preferred embodiments of the invention, the BLGisolate powder has a degree of protein denaturation of at least 11%,preferably at least 20%, more preferably at least 40%, even morepreferably at least 50%, even more preferably at least 75%, and mostpreferably at least 90%.

If BLG isolate powder has a significant level of protein denaturation itis often preferred to keep a low level of insoluble protein matter, i.e.precipitated protein matter that would settle in a beverage duringstorage. The level of insoluble matter is measure according to Example1.10.

In some preferred embodiments of the invention the BLG isolate powdercomprises at most 20% w/w insoluble protein matter, preferably at most10% w/w insoluble protein matter, more preferably at most 5% w/winsoluble protein matter, even more preferred at most 3% w/w insolubleprotein matter, and most preferred at most 1% w/w insoluble proteinmatter. It may even be preferred that the BLG isolate powder does notcontain any insoluble protein matter at all.

In some preferred embodiments of the invention the BLG isolate powderhas a crystallinity of BLG of at most 19%, preferably at most 10%, morepreferably at most 5%, and most preferably 0%.

In other preferred embodiments of the invention the BLG isolate powderhas a crystallinity of BLG of at least 20%, preferably at least 40%,more preferably at least 60%, and most preferably at least 80%. Theseembodiments contain a significant amount of dried BLG crystals andprovide the benefits of having the protein source present in solid,compact form.

The present inventors have found that the heat-stability at pH 3.9 of aBLG isolate powder is a good indicator for its usefulness fortransparent high protein beverages. The heat-stability at pH 3.9 ismeasured according to Example 1.2.

It is particularly preferred that the BLG isolate powder has aheat-stability at pH 3.9 of at most 200 NTU, preferably at most 100 NTU,more preferred at most 60 NTU, even more preferred at most 40 NTU, andmost preferred at most 20 NTU. Even better heat-stabilities are possibleand the BLG isolate powder preferably has a heat-stability at pH 3.9 ofat most 10 NTU, preferably at most 8 NTU, more preferred at most 4 NTU,even more preferred at most 2 NTU.

The content of microorganisms of the BLG isolate powder is preferablykept to a minimum. However, it is a challenge to obtain both a highdegree of protein nativeness and a low content of microorganism asprocesses for microbial reduction tend to lead to protein unfolding anddenaturation. The present invention makes it possible to obtain a verylow content of microorganism while at the same time maintain a highlevel of the nativeness of BLG.

Thus, in some preferred embodiments of the invention, the BLG isolatepowder contains at most 15000 colony-forming units (CFU)/g. Preferably,the BLG isolate powder contains at most 10000 CFU/g. More preferably,the BLG isolate powder contains at most 5000 CFU/g. Even morepreferably, the BLG isolate powder contains at most 1000 CFU/g. Evenmore preferably, the BLG isolate powder contains at most 300 CFU/g. Mostpreferably, the BLG isolate powder contains at most 100 CFU/g such ase.g. at most 10 CFU/g. In a particularly preferred embodiment the powderis sterile. A sterile BLG isolate powder may e.g. be prepared bycombining several physical microbial reduction processes during theproduction of the BLG isolate powder, such as e.g. microfiltration andheat-treatment at acidic pH.

In some preferred embodiments of the invention, the BLG isolate powderhas a pH in the range of i) 2-4.9, ii) 6.1-8.5, or iii) 5.0-6.0 andcomprises:

-   -   total protein in an amount of at least 30% w/w, preferably at        least 80% w/w, and even more preferably at least 90% w/w    -   beta-lactoglobulin (BLG) in an amount of at least 85% w/w        relative to total protein, preferably at least 90% w/w,    -   water in an amount of at most 6% w/w,    -   lipid in an amount of at most 2% w/w, preferably at most 0.5%        w/w,        said BLG isolate powder having:    -   an intrinsic tryptophan fluorescence emission ratio (I330/I350)        of at least 1.11,    -   a degree of protein denaturation of at most 10%, and    -   a heat-stability at pH 3.9 of at most 200 NTU.

In some preferred embodiments of the invention, the BLG isolate powderhas a pH in the range of i) 2-4.9 or ii) 6.1-8.5 and comprises:

-   -   total protein in an amount of at least 30% w/w, preferably at        least 80% w/w, and even more preferably at least 90% w/w    -   beta-lactoglobulin (BLG) in an amount of at least 85% w/w        relative to total protein, preferably at least 90% w/w, and more        preferably at least 94% w/w relative to total protein    -   water in an amount of at most 6% w/w,    -   lipid in an amount of at most 2% w/w, preferably at most 0.5%        w/w,

said BLG isolate powder having:

-   -   an intrinsic tryptophan fluorescence emission ratio (I330/I350)        of at least 1.11,    -   a degree of protein denaturation of at most 10%, preferably at        most 5%, and    -   a heat-stability at pH 3.9 of at most 70 NTU, preferably at most        50 NTU and even more preferably at most 40 NTU.

In some preferred embodiments of the invention the BLG isolate powderhas a pH in the range of i) 2-4.9 or ii) 6.1-8.5 and comprises:

-   -   total protein in an amount of at least 30% w/w,    -   beta-lactoglobulin (BLG) in an amount of at least 85% w/w        relative to total protein, preferably at least 90% w/w,    -   water in an amount of at most 6% w/w,        said BLG isolate powder having:    -   a bulk density of at least 0.2 g/cm³,    -   an intrinsic tryptophan fluorescence emission ratio (I330/I350)        of at least 1.11,    -   a degree of protein denaturation of at most 10%, and    -   a heat-stability at pH 3.9 of at most 200 NTU.

In other preferred embodiments of the invention, the BLG isolate powderhas a pH in the range of 2-4.9 and comprises:

-   -   total protein in an amount of at least 80% w/w, preferably at        least 90% w/w, and even more preferably at least 94% w/w    -   beta-lactoglobulin (BLG) in an amount of at least 85% w/w        relative to total protein, preferably at least 90% w/w, and even        more preferably at least 94% w/w relative to total protein,    -   water in an amount of at most 6% w/w,    -   lipid in an amount of at most 2% w/w, preferably at most 0.5%        w/w,        said BLG isolate powder having:    -   a bulk density of at least 0.2 g/cm³, preferably at least 0.3        g/cm³, and more preferably at least 0.4 g/cm³,    -   an intrinsic tryptophan fluorescence emission ratio (I330/I350)        of at least 1.11,    -   a degree of protein denaturation of at most 10%, preferably at        most 5%, and more preferably at most 2%, and    -   a heat-stability at pH 3.9 of at most 50 NTU, preferably at most        30 NTU and even more preferably at most 10 NTU.

In yet other preferred embodiments of the invention, the BLG isolatepowder has a pH in the range of 6.1-8.5 and comprises:

-   -   total protein in an amount of at least 80% w/w, preferably at        least 90% w/w, and even more preferably at least 94% w/w    -   beta-lactoglobulin (BLG) in an amount of at least 85% w/w        relative to total protein, preferably at least 90% w/w, and even        more preferably at least 94% w/w relative to total protein,    -   water in an amount of at most 6% w/w,    -   lipid in an amount of at most 2% w/w, preferably at most 0.5%        w/w,        said BLG isolate powder having:    -   a bulk density of at least 0.2 g/cm³, preferably at least 0.3        g/cm³, and more preferably at least 0.4 g/cm³,    -   a degree of protein denaturation of at most 10%, preferably at        most 5%, and more preferably at most 2%, and    -   a heat-stability at pH 3.9 of at most 50 NTU, preferably at most        30 NTU, and even more preferably at most 10 NTU.

In further preferred embodiments of the invention, the BLG isolatepowder has a pH in the range of 6.1-8.5 and comprises:

-   -   total protein in an amount of at least 80% w/w, preferably at        least 90% w/w, and even more preferably at least 94% w/w    -   beta-lactoglobulin (BLG) in an amount of at least 85% w/w        relative to total protein, preferably at least 90% w/w, and even        more preferably at least 94% w/w relative to total protein,    -   water in an amount of at most 6% w/w,    -   lipid in an amount of at most 2% w/w, preferably at most 0.5%        w/w,        said BLG isolate powder having:    -   a bulk density of at least 0.2 g/cm³, preferably at least 0.3        g/cm³, and more preferably at least 0.4 g/cm³,    -   a degree of protein denaturation of at most 10%, preferably at        most 5%, and more preferably at most 2%, and    -   a heat-stability at pH 3.9 of at most 50 NTU, preferably at most        30 NTU, and even more preferably at most 10 NTU.

In further preferred embodiments of the invention, the BLG isolatepowder has a pH in the range of 5.0-6.0 and comprises:

-   -   total protein in an amount of at least 80% w/w, preferably at        least 90% w/w, and even more preferably at least 94% w/w,    -   beta-lactoglobulin (BLG) in an amount of at least 85% w/w        relative to total protein, preferably at least 90% w/w, and even        more preferably at least 94% w/w relative to total protein,    -   water in an amount of at most 6% w/w,    -   lipid in an amount of at most 2% w/w, preferably at most 0.5%        w/w,        said BLG isolate powder having:    -   a bulk density of at least 0.2 g/cm³, preferably at least 0.3        g/cm³, and more preferably at least 0.4 g/cm³,    -   a degree of protein denaturation of at most 10%, preferably at        most 5%, and more preferably at most 2%,    -   a heat-stability at pH 3.9 of at most 50 NTU, preferably at most        30 NTU, and even more preferably at most 10 NTU, and    -   preferably, a BLG crystallinity of less than 10%.

The BLG isolate powder containing BLG in an amount of at least 85% w/wrelative to total protein, is typically provided by a method comprisingthe steps of:

a) providing a liquid BLG isolate having

-   -   i) a pH in the range of 2-4.9,    -   ii) a pH of in the range of 6.1-8.5, or    -   iii) a pH of in the range of 5.0-6.0    -   said liquid BLG isolate containing BLG in an amount of at least        85 w/w relative to total protein,        b) optionally, subjecting the liquid BLG isolate to a physical        microbial reduction,        c) drying the liquid BLG isolate, preferably by spray-drying.

The BLG isolate is preferably prepared from mammal milk, and preferablyfrom ruminant milk such as e.g. milk from cow, sheep, goat, buffalo,camel, llama, mare and/or deer. Protein derived from bovine milk isparticularly preferred. The BLG is therefore preferably bovine BLG.

The liquid BLG isolate may be provided in a number of different ways.

Typically, the provision of the liquid BLG isolate involves, or evenconsists of, isolating BLG from a whey protein feed to provide aBLG-enriched composition by one or more of the following methods:

-   -   crystallisation or precipitation of BLG by salting-in,    -   crystallisation or precipitation of BLG of BLG by salting-out,    -   ion exchange chromatography, and    -   fractionation of whey proteins by ultrafiltration.

A particularly preferred way of providing the BLG-enriched compositionis by crystallisation of BLG, preferably by salting-in or alternativelyby salting-out.

The whey protein feed is preferably a WPC, a WPI, an SPC, an SPI, or acombination thereof.

The term “whey protein feed” pertains to the composition from which theBLG-enriched composition and subsequently the liquid BLG isolate arederived.

In some embodiments of the invention, the preparation of theBLG-enriched composition includes, or even consist of, high salt BLGcrystallisation in the pH range 3.6-4.0 according to U.S. Pat. No.2,790,790 A1.

In other embodiments of the invention the preparation of theBLG-enriched composition includes, or even consists of, the methoddescribed by de Jongh et al (Mild Isolation Procedure Discloses NewProtein Structural Properties of β-Lactoglobulin, J Dairy Sci., vol.84(3), 2001, pages 562-571) or by Vyas et al (Scale-Up of Nativeβ-Lactoglobulin Affinity Separation Process, 3. Dairy Sci. 85:1639-1645,2002).

However, in particularly preferred embodiments of the invention, theBLG-enriched composition is prepared by crystallisation at pH 5-6 undersalting-in conditions as described in the PCT applicationPCT/EP2017/084553, which is incorporated herein by reference for allpurposes.

In some preferred embodiments of the invention, the BLG-enrichedcomposition is an edible BLG composition according to PCT/EP2017/084553containing at least 90% BLG relative to total protein and preferablycontaining BLG crystals.

If it does not already have the required characteristics to be used asliquid BLG isolate, the BLG-enriched composition which has been isolatedfrom whey protein feed may be subjected to one or more steps selectedfrom the group of:

-   -   demineralisation,    -   addition of minerals    -   dilution,    -   concentration,    -   physical microbioal reduction, and    -   pH adjustment        as part of providing the liquid BLG isolate.

Non-limiting examples of demineralisation include e.g. dialysis, gelfiltration, UF/diafiltration, NF/diafiltration, and ion exchangechromatography.

Non-limiting examples of addition of minerals include addition ofsoluble, food acceptable salts, such as e.g. salts of Na, K, Ca, and/orMg. Such salts may e.g. be phosphate-salts, chloride salts or salts offood acids, such as e.g. citrate salt or lactate salt. The minerals maybe added in solid, suspended, or dissolved form.

Non-limiting examples of dilution include e.g. addition of liquiddiluent such as water, demineralised water, or aqueous solutions ofminerals, acids or bases.

Non-limiting examples of concentration include e.g. evaporation, reverseosmosis, nanofiltration, ultrafiltration and combinations thereof.

If the concentration has to increase the concentration of proteinrelative to total solids, it is preferred to use concentration stepssuch as ultrafiltration or alternatively dialysis. If the concentrationdoes not have to increase the concentration of protein relative to totalsolids, methods such as e.g. evaporation, nanofiltration and/or reverseosmosis can be useful.

Non-limiting examples of physical microbioal reduction include e.g.heat-treatment, germ filtration, UV radiation, high pressure treatment,pulsed electric field treatment, and ultrasound. These methods arewell-known to the person skilled in the art.

Non-limiting examples of pH adjustment include e.g. addition of basesand/or acids, and preferably food acceptable bases and/or acids. It isparticularly preferred to employ acids and/or bases that are capable ofchelating divalent metal cations. Examples of such acids and/or basesare citric acid, citrate salt, EDTA, lactic acid, lactate salt,phosphoric acid, phosphate salt, and combinations thereof.

In the following a number of preferred embodiments of the provision ofthe liquid BLG isolate of step a) from BLG-enriched composition. Theprocess steps mentioned in this context are applied to theBLG-containing product stream following the BLG-enriched composition.

The present invention has been described above with reference tospecific embodiments. However, other embodiments than the abovedescribed are equally possible within the scope of the invention. Thedifferent features and steps of various embodiments and aspects of theinvention may be combined in other ways than those described hereinunless it is stated otherwise.

EXAMPLES Example 1: Methods of Analysis Example 1.1: Determination ofProtein Nativeness by Intrinsic Tryptophan Fluorescence

Tryptophan (Trp) fluorescence spectroscopy is a well-described tool tomonitor protein folding and unfolding. Trp residues buried within nativeproteins typically display highest fluorescence emission around 330 nmthan when present in more solvent exposed positions such as unfoldedproteins. In unfolded proteins, the wavelengths for Trp fluorescenceemission typically shift to higher wavelengths and are often measuredaround 350 nm. We here exploit this transition to monitor thermallyinduced unfolding by calculating the ratio between fluorescence emissionat 330 nm and 350 nm to investigate the influence of heatingtemperature.

The analysis comprises the following steps:

-   -   Beverage compositions were diluted to 0.6 mg/ml in MQ water.    -   300 μl sample was transferred to white 96-well plate avoiding        bubbles or 3 mL was transferred to 10 mm quartz cuvette.    -   The tryptophan fluorescence emission intensity between 310 and        400 nm was recorded from the top by excitation at 295 using 5 nm        slits.    -   Samples were measured at 22 degrees C. using a Cary Eclipse        fluorescence spectrophotometer equipped with a plate reader        accessory (G9810A) or single cuvette holder.    -   The emission intensity ratio was calculated by dividing the        measured fluorescence emission intensity at 330 nm with the        emission intensity at 350 nm, R=I330/I350, and used as a measure        of protein nativity.        -   R of at least 1.11 describes a predominant native BLG            conformation and        -   R of less than 1.11 reports on at least partial unfolding            and aggregation.

Example 1.2: Heat-Stability at pH 3.9 Heat-Stability at pH 3.9:

The heat-stability at pH 3.9 is a measure of the ability of proteincomposition to stay clear upon prolonged pasteurization at pH 3.9.

The heat-stability at pH 3.9 is determined by forming an aqueoussolution having a pH of 3.9 and comprising 6.0% w/w protein by mixing asample of the powder or liquid to be tested with water (or alternativelyconcentrating it by low temperature evaporation if it is a diluteliquid) and adjusting the pH to 3.9 with the minimum amount of 0.1 MNaOH or 0.1 M HCl required.

The pH-adjusted mixture is allowed to rest for 30 minutes after which 25mL of the mixture is transferred to a 30 mL thin-walled glass test tube.It is heated to 75.0 degrees C. for 300 seconds by immersion into awater-bath having a temperature of 75.0 degrees C. Immediately after theheating, the glass test tube is cooled to 1-5 degrees C. by transferringit to an ice bath and the turbidity of the heat-treated sample ismeasured according to Example 1.7.

Example 1.3: Determination of the Degree of Protein Denaturation of aWhey Protein Composition

Denatured whey protein is known to have a lower solubility at pH 4.6than at pH values below or above pH 4.6, therefore the degree ofdenaturation of a whey protein composition is determined by measuringthe amount of soluble protein at pH 4.6 relative to the total amount ofprotein at a pH where the proteins in the solution are stable.

More specifically for whey proteins, the whey protein composition to beanalysed (e.g. a powder or an aqueous solution) is converted to:

-   -   a first aqueous solution containing 5.0% (w/w) total protein and        having a pH of 7.0 or 3.0, and    -   a second aqueous solution containing 5.0% (w/w) total protein        and having a pH of 4.6.

pH adjustments are made using 3% (w/w) NaOH (aq) or 5% (w/w) HCl (aq).

The total protein content (P_(pH 7.0 or 3.0)) of the first aqueoussolution is determined according to example 1.5.

The second aqueous solution is stored for 2 h at room temperature andsubsequently centrifuged at 3000 g for 5 minutes. A sample of thesupernatant is recovered and analysed according to Example 1.5 to givethe protein concentration in the supernatant (S_(pH4.6)).

The degree of protein denaturation, D, of the whey protein compositionis calculated as:

D=((P _(pH 7.0 or 3.0) −S _(pH 4.6))/P _(pH 7.0 or 3.0))*100%

Example 1.4 Determination of Protein Denaturation (with pH 4.6 AcidPrecipitation) Using Reverse Phase UPLC Analysis

BLG samples (such as non-heated reference and heated BLG beveragecompositions) were diluted to 2% in MQ water. 5 mL protein solution, 10mL Milli-Q, 4 mL 10% acetic acid and 6 mL 1.0M NaOAc are mixed andstirred for 20 minutes to allow precipitation agglomeration of denaturedprotein around pH 4.6. The solution is filtered through 0.22 μm filterto remove agglomerates and non-native proteins.

All samples were subjected to the same degree of dilution by addingpolished water.

For each sample, the same volume was loaded on an UPLC system with aUPLC column (Protein BEH C4; 300 Å; 1.7 μm; 150×2.1 mm) and detected at214 nm.

The samples were run using the following conditions:

Buffer A: Milli-Q water, 0.1% w/w TFABuffer B: HPLC grade acetonitrile, 0.1% w/w TFAFlow: 0.4 ml/minGradient: 0-6.00 minutes 24-45% B; 6.00-6.50 minutes 45-90% B; 6.50-7.00minutes 90% B; 7.00-7.50 minutes 90-24% B and 7.50-10.00 minutes 24% B.

The area of BLG peaks against a protein standard (Sigma L0130) was usedto determine the concentration of native bLG in samples (5 levelcalibration curve)

Samples were diluted further and reinjected if outside linear range.

Example 1.5: Determination Total Protein

The total protein content (true protein) of a sample is determined by:

1) Determining the total nitrogen of the sample following ISO8968-1/2|IDF 020-1/2—Milk—Determination of nitrogen content—Part 1/2:Determination of nitrogen content using the Kjeldahl method.2) Determining the non-protein nitrogen of the sample following ISO8968-4|IDF 020-4—Milk—Determination of nitrogen content—Part 4:Determination of non-protein-nitrogen content.3) Calculating the total amount protein as(m_(total nitrogen)−m_(non-protein-nitrogen))*6.38.

Example 1.6: Determination of Non-Aggregated BLG, ALA, and CMP

The content of non-aggregated alpha-lactalbumin (ALA),beta-lactoglobulin (BLG) and caseinomacropeptide (CMP), respectively wasanalysed by HPLC analysis at 0.4 mL/min. 25 microL filtered sample isinjected onto 2 TSKgel3000PWxl (7.8 mm 30 cm, Tosohass, Japan) columnsconnected in series with attached pre-column PWxl (6 mm×4 cm, Tosohass,Japan) equilibrated in the eluent (consisting of 465 g Milli-Q water,417.3 g acetonitrile and 1 mL triflouroacetic acid) and using a UVdetector at 210 nm.

Quantitative determination of the contents of native alpha-lactalbumin(C_(alpha)), beta-lactoglobulin (C_(beta)), and caseinomacropeptide(C_(CMP)) was performed by comparing the peak areas obtained for thecorresponding standard proteins with those of the samples.

The total amount of additional protein (non-BLG protein) was determinedby subtracting the amount of BLG from the amount of total protein(determined according to Example 1.5)

Example 1.7: Determination of Turbidity

Turbidity is the cloudiness or haziness of a fluid caused by largenumber of particles that are generally invisible to the naked eye,similar to smoke in air.

Turbidity is measured in nephelometric turbidity units (NTU).

20 mL beverages/samples were added to NTU-glass and placed in theTurbiquant® 3000 IR Turbidimeter. The NTU-value was measured afterstabilisation and repeated twice.

Example 1.8: Determination of Viscosity

The viscosity of beverage preparations was measured using a Rheometer(Anton Paar, Physica MCR301).

3.8 mL sample was added to cup DG26.7. Samples were equilibrated to 22°C., then pre-sheared for 30 sec. at 50 s⁻¹, followed by a 30 sec.equilibrium time and shear rate sweeps between 1 s⁻¹ and 200 s⁻¹ and 1s⁻¹ were performed.

The viscosity is presented in the unit centipoise (cP) at a shear rateof 100 s⁻¹ unless otherwise stated. The higher the measured cP values,the higher the viscosity.

Alternatively, the viscosity was estimated using a Viscoman by Gilsonand reported at a shear rate of about 300 s⁻¹

Example 1.9: Determination of Colour

The colour was measured using a Chroma Meter (Konica Minolta, CR-400).15 g sample was added to a small petri dish (55×14.2 mm, VWR Cat#391-0895) avoiding bubble formation. The protein content of the sampleswas standardised to 6.0 w/w % protein or less.

The Chroma Meter was calibrated to a white calibration plate (No.19033177). The illuminant was set to D65 and the observer to 2 degree.The color (CIELAB color space, a*-, b*-, L*-value) was measured withlids covering the suspension, as the average of three individualreadings in different places of the petri dish.

Demineralised water reference has the following values:

L* 39.97±0.3

a* 0.00±0.06b* −0.22±0.09

The measurements were converted to delta/difference values based ondemineralised water measurement.

delta L*=L _(sample standardised to 6.0 w/w % protein) *−L_(demin. water)*, measured at room temperature.

delta a*=a _(sample standardised to 6.0 w/w % protein) *−a_(demin. water)*, measured at room temperature.

delta b*=b _(sample standardised to 6.0 w/w % protein) *−b_(demin. water)*, measured at room temperature.

The samples is standardized to 6.0 w/w % protein or below.

The L*a*b* colour space (also referred to as the CIELAB space) is one ofthe uniform colour spaces defined by the International Commission onIllumination (CIE) in 1976 and was used to quantitatively reportlightness and hue (ISO 11664-4:2008(E)/CIE S 014-4/E:2007).

In this space, L* indicates lightness (value from 0-100), the darkestblack at L*=0, and the brightest white at L*=100.

The colour channels a* and b*, represent true neutral grey values ata*=0 and b*=0. The a* axis represents the green-red component, withgreen in the negative direction and red in the positive direction. Theb* axis represents the blue-yellow component, with blue in the negativedirection and yellow in the positive direction.

Example 1.10 Beverage Stability Test/Insoluble Protein Matter

Whey protein beverage compositions were considered stable if less than15% of total protein in heated samples precipitated upon centrifugationat 3000 g for 5 minutes:

-   -   Approx. 20 g samples were added to centrifuge tubes and        centrifugated at 3000 g 5 min.    -   Kjeldahl analysis of protein before centrifugation and the        supernatant after centrifugation were used to quantify protein        recovery See example 1.5

The loss of protein is calculated:

${{Denatu}\;{ration}\mspace{14mu}\%} = {\left( \frac{P_{total} - P_{3000{xg}}}{P_{total}} \right)*100\%}$

This parameter is also sometimes referred to as the level of insolubleprotein matter and can be used for analyzing both liquid and powdersamples. If the sample is a powder, 10 g of the powder is suspended in90 g demineralized water and allowed to hydrate at 22 degrees C. undergentle stirring for 1 hours. Approx. 20 g of sample (e.g. liquid sampleor the suspended powder sample) to centrifuge tubes and centrifugated at3000 g 5 min. Kjeldahl analysis of protein before centrifugation(P_(total)) and the supernatant after centrifugation (P_(3000×g)) wereused to quantify protein recovery according to Example 1.5.

The amount of insoluble protein matter is calculated:

${{per}{cen}}\;{tage}\mspace{14mu}{of}\mspace{14mu}{insoluble}\mspace{14mu}{protein}\mspace{14mu}{matter}{= {\left( \frac{P_{total} - P_{3000{xg}}}{P_{total}} \right)*100\%}}$

Example 1.11: Sensory Evaluation

The heat-treated beverage preparations underwent a descriptive sensoryevaluation. The beverage preparations had been subjected to heat usingplate heat exchangers.

1 volume sample was mixed with 1 volume water and compared to non-heatedwhey protein isolate, lactic acid and citric acid are also used to forman attribute list prior to the final tasting session:

Category Attributes: Aroma Whey, acidic (sour milk product) Basic tasteAcid, bitter Flavour Whey, citric acid, lactic acid Mouth feelingDrying, astringency

Crackers, white tea, melon and water were used to cleanse the mouth ofparticipants between each sample.

15 mL test sample at ambient temperature (20-25 degrees C.) was servedin small cups.

Test samples were each served to 10 individuals three times in threedifferent blocks in randomised order.

The attributes (see table above) were rated on a 15 cm scale with 0=lowintensity and 15=high intensity.

The statistical analysis was conducted in ‘Panelcheck’ software using a3-way ANOVA test for multiple replicates. Samples were fixed and panelwas set to random.

Bonferroni correction implying least significance difference values(pairwise comparisons of groups associated to a letter) was used toevaluate significant differences between samples.

Example 1.12: Determination of Transparency by Imaging

Photographs of beverage preparations were conducted by placing samplesin turbidity NTU measuring vials touching a piece of paper with ‘loremipsum’ text. Vials were photographed using a smartphone and theinventors evaluated whether the text could be clearly observed throughthe vial.

Example 1.13: Determination of Ash Content

The ash content of a food product is determined according to NMKL173:2005 “Ash, gravimetric determination in foods”.

Example 1.14: Determination of Conductivity

The “conductivity” (sometimes referred to as the “specific conductance”)of an aqueous solution is a measure of the ability of the solution toconduct electricity. The conductivity may e.g. be determined bymeasuring the AC resistance of the solution between two electrodes andthe result is typically given in the unit milliSiemens per cm (mS/cm).The conductivity may for example be measured according to the EPA (theUS Environmental Protection Agency) Method No. 120.1

Conductivity values mentioned herein have been normalised to 25 degreesC. unless it is specified otherwise.

The conductivity is measured on a Conductivity meter (WTW Cond 3210 witha tetracon 325 electrode).

The system is calibrated as described in the manual before use. Theelectrode is rinsed thoroughly in the same type of medium as themeasurement is conducted on, in order to avoid local dilutions. Theelectrode is lowered into the medium so that the area where themeasurement occurs is completely submerged. The electrode is thenagitated so that any air trapped on the electrode is removed. Theelectrode is then kept still until a stable value can be obtained andrecorded from the display.

Example 1.15: Determination of the Total Solids of a Solution

The total solids of a solution may be determined according NMKL 1102^(nd) Edition, 2005 (Total solids (Water)-Gravimetric determination inmilk and milk products). NMKL is an abbreviation for “NordiskMetodikkomité for Næringsmidler”.

The water content of the solution can be calculated as 100% minus therelative amount of total solids (% w/w).

Example 1.16: Determination of pH

All pH values are measured using a pH glass electrode and are normalisedto 25 degrees C. The pH glass electrode (having temperaturecompensation) is rinsed carefully before and calibrated before use.

When the sample is in liquid form, then pH is measured directly in theliquid solution at 25 degrees C.

When the sample is a powder, 10 gram of a powder is dissolved in 90 mlof demineralised water at room temperature while stirring vigorously.The pH of the solution is then measured at 25 degrees C.

Example 1.17: Determination of Loose Density and Bulk Density

The density of a dry powder is defined as the relation between weightand volume of the powder which is analysed using a special Stampfvolumeter (i.e. a measuring cylinder) under specified conditions. Thedensity is typically expressed in g/ml or kg/L.

In this method, a sample of dried powder is tamped in a measuringcylinder. After a specified number of tappings, the volume of theproduct is read and the density is calculated.

Three types of densities can be defined by this method:

-   -   Poured density, which is the mass divided with the volume of        powder after it has been transferred to the specified measuring        cylinder.    -   Loose density, which is the mass divided with the volume of        powder after 100 tappings according to the specified conditions        in this standard.    -   Bulk density, which is the mass divided with the volume of        powder after 625 tappings according to the specified conditions        in this standard.

The method uses a special measuring cylinder, 250 ml, graduated 0-250ml, weight 190±15 g (3. Engelsmann A. G. 67059 Ludwigshafen/Rh) and aStampf volumeter, e.g. 3. Engelsmann A. G.

The loose density and the bulk density of the dried product aredetermined by the following procedure.

Pre-Treatment:

The sample to be measured is stored at room temperature.

The sample is then thoroughly mixed by repeatedly rotating and turningthe container (avoid crushing particles). The container is not filledmore than ⅔.

Procedure:

Weigh 100.0±0.1 gram of powder and transfer it to the measuringcylinder. The volume V₀ is read in ml.

If 100 g powder does not fit into the cylinder, the amount should bereduced to 50 or 25 gram.

Fix the measuring cylinder to the Stampf volumeter and let it tap 100taps. Level the surface with the spatula and read the volume V₁₀₀ in ml.

Change the number of tabs to 625 (incl. the 100 taps). After tapping,level the surface and read the volume V₆₂₅ in ml.

Calculation of Densities:

Calculate the loose and the bulk densities expressed in g/ml accordingto the following formula:

Bulk density=M/V

where M designates weighed sample in grams and V designates volume after625 tappings in ml.

Example 1.18: Determination of the Water Content of a Powder

The water content of a food product is determined according to ISO5537:2004 (Dried milk—Determination of moisture content (Referencemethod)). NMKL is an abbreviation for “Nordisk Metodikkomité forNæringsmidler”.

Example 1.19: Determination of the Amounts of Calcium, Magnesium,Sodium, Potassium, Phosphorus (ICP-MS Method)

The total amounts of calcium, magnesium, sodium, potassium, andphosphorus are determined using a procedure in which the samples arefirst decomposed using microwave digestion, and then the total amount ofmineral(s) is determined using an ICP apparatus.

Apparatus:

The microwave is from Anton Paar and the ICP is an Optima 2000DV fromPerkinElmer Inc.

Materials: 1 M HNO₃ Yttrium in 2% HNO₃

Suitable standards for calcium, magnesium, sodium, potassium, andphosphorus in 5% HNO₃

Pre-Treatment:

Weigh out a certain amount of powder and transfer the powder to amicrowave digestion tube. Add 5 mL 1M HNO₃. Digest the samples in themicrowave in accordance with microwave instructions. Place the digestedtubes in a fume cupboard, remove the lid and let volatile fumesevaporate.

Measurement Procedure:

Transfer pre-treated sample to DigiTUBE using a known amount of Milli-Qwater. Add a solution of yttrium in 2% HNO₃ to the digestion tube (about0.25 mL per 50 mL diluted sample) and dilute to known volume usingMilli-Q water. Analyse the samples on the ICP using the proceduredescribed by the manufacturer.

A blind sample is prepared by diluting a mixture of 10 mL 1M HNO₃ and0.5 mL solution of yttrium in 2% HNO₃ to a final volume of 100 mL usingMilli-Q water.

At least 3 standard samples are prepared having concentrations whichbracket the expected sample concentrations.

Example 1.20: Determination of the Furosine-Value

The furosine value is determined as described in “Maillard ReactionEvaluation by Furosine Determination During Infant Cereal Processing”,Guerra-Hernandez et al, Journal of Cereal Science 29 (1999) 171-176, andthe total amount of protein is determined according to Example 1.5. Thefurosine value is reported in the unit mg furosine per 100 g protein.

Example 1.21: Determination of the Crystallinity of BLG in a Liquid

The following method is used to determine the crystallinity of BLG in aliquid having a pH in the range of 5-6.

a) Transfer a 10 mL sample of the liquid in question to a Maxi-Spinfilter with a 0.45 micron pore size CA membrane.b) Immediately spin the filter at 1500 g for 5 min. keeping thecentrifuge at 2 degrees C.c) Add 2 mL cold Milli-Q water (2 degrees C.) to the retentate side ofthe spin filter and immediately, spin the filter at 1500 g for 5 minwhile keeping the centrifuge cooled at 2 degrees C., collect thepermeate (permeate A), measure the volume and determine BLGconcentration via HPLC using the method outlined in Example 1.31.d) Add 4 mL 2M NaCl to the retentate side of the filter, agitate quicklyand allow the mixture to stand for 15 minutes at 25 degrees C.e) Immediately spin the filter at 1500 g for 5 min and collect thepermeate (permeate B)f) Determine the total weight of BLG in permeate A and permeate B usingthe method outlined in Example 1.31 and convert the results to totalweight of BLG instead of weight percent. The weight of BLG in permeate Ais referred to as m_(Permeate A) and the weight of BLG in permeate B isreferred to as m_(permeate B).

-   -   g) The crystallinity of the liquid with respect to BLG is        determined as:

crystallinity=m _(Permeate B)/(m _(permeate A) +m _(Permeate B))*100%

Example 1.22: Determination of the Crystallinity of BLG in a Dry Powder

This method is used to determine the crystallinity of BLG in a drypowder.

a) 5.0 gram of the powder sample is mixed with 20.0 gram of cold Milli-Qwater (2 degrees C.) and allowed to stand for 5 minute at 2 degrees C.b) Transfer the sample of the liquid in question to a Maxi-Spin filterwith a 0.45 micron CA membrane.c) Immediately spin the filter at 1500 g for 5 min. keeping thecentrifuge at 2 degrees C.d) Add 2 mL cold Milli-Q water (2 degrees C.) to the retentate side ofthe spin filter and immediately, spin the filter at 1500 g for 5 min,collect the permeate (permeate A), measure the volume and determine BLGconcentration via HPLC using the method outlined in Example 1.6 andconvert the results to total weight of BLG instead of weight percent.The weight of BLG in permeate A is referred to as m_(permeate A)f) The crystallinity of BLG in the powder is then calculated using thefollowing formula:

${crystallinity} = {\frac{m_{{BLG}\mspace{11mu}{total}} - m_{{permeate}\mspace{11mu} A}}{m_{{BLG}\mspace{11mu}{total}}}*100\%}$

where m_(BLG total) is the total amount of BLG in the powder sample ofstep a).

If the total amount of BLG of powder sample is unknown, this may bedetermined by suspending another 5 g powder sample (from the same powdersource) in 20.0 gram of Milli-Q water, adjusting the pH to 7.0 byaddition of aqueous NaOH, allowing the mixture to stand for 1 hour at 25degrees C. under stirring, and finally determining the total amount ofBLG of the powder sample using Example 1.31.

Example 1.23: Determination of UF Permeate Conductivity

15 mL of sample is transferred to an Amicon Ultra-15 Centrifugal FilterUnits with a 3 kDa cut off (3000 NMWL) and centrifugated at 4000 g for20-30 minutes or until a sufficient volume of UF permeate for measuringconductivity is accumulated in the bottom part of the filter units. Theconductivity is measured immediately after centrifugation. The samplehandling and centrifugation are performed at the temperature of thesource of the sample.

Example 1.24: Detection of Dried BLG Crystals in a Powder

The presence of dried BLG crystals in a powder can be identified thefollowing way:

A sample of the powder to be analysed is re-suspended and gently mixedin demineralised water having a temperature of 4 degrees C. in a weightratio of 2 parts water to 1 part powder, and allowed to rehydrate for 1hour at 4 degrees C.

The rehydrated sample is inspected by microscopy to identify presence ofcrystals, preferably using plan polarised light to detect birefringence.

Crystal-like matter is separated and subjected to x-ray crystallographyin order verify the existence of crystal structure, and preferably alsoverifying that the crystal lattice (space group and unit celldimensions) corresponds to those of a BLG crystal.

The chemical composition of the separated crystal-like matter isanalysed to verify that its solids primarily consists of BLG.

Example 1.25: Determination of the Total Amount of Lactose

The total amount of lactose is determined according to ISO 5765-2:2002(IDF 79-2: 2002) “Dried milk, dried ice-mixes and processedcheese—Determination of lactose content—Part 2: Enzymatic methodutilizing the galactose moiety of the lactose”.

Example 1.26: Determination of the Total Amount of Carbohydrate

The amount of carbohydrate is determined by use of Sigma Aldrich TotalCarbohydrate Assay Kit (Cat MAK104-1KT) in which carbohydrates arehydrolysed and converted to furfural and hydroxyfurfurals which areconverted to a chromagen that is monitored spectrophotometrically at 490nm.

Example 1.27: Determination of the Total Amount of Lipids

The amount of lipid is determined according to ISO 1211:2010(Determination of Fat Content—Röse-Gottlieb Gravimetric Method).

Example 1.28: Determination of Brix

Brix measurements were conducted using a PAL-α digital hand-heldrefractometer (Atago) calibrated against polished water (water filteredby reverse osmosis to obtain a conductivity of at most 0.05 mS/cm).

Approx. 500 μl of sample was transferred to the prism surface of theinstrument and the measurement was started. The measured value was readand recorded

The Brix of a whey protein solution is proportional to the content oftotal solids (TS) and TS (% w/w) is approx. Brix*0.85.

Example 1.29 Determination of Lactoferrin and Lactoperoxidase

The concentration of lactoferrin is determined by an ELISA immunoassayas outlined by Soyeurt 2012 (Soyeurt et al; Mid-infrared prediction oflactoferrin content in bovine milk: potential indicator of mastitis;Animal (2012), 6:11, pp 1830-1838)

The concentration of lactoperoxidase is determined using a commerciallyavailable bovine lactoperoxidase kit.

Example 1.30: Determination the Number of Colony-Forming Units

The determination of the number of colony-forming units per gram sampleis performed according to ISO 4833-1:2013(E): Microbiology of food andanimal feeding stuffs—horizontal method for the enumeration ofmicroorganisms—Colony-count technique at 30° C.

Example 1.31: Determination of the Total Amount of BLG, ALA, and CMP

This procedure is a liquid chromatographic (HPLC) method for thequantitative analysis of proteins such as ALA, BLG and CMP andoptionally also other protein species in a composition.

Contrary to the method of Example 1.6 the present method also measuresproteins that are present in aggregated and therefore provides a measureof the total amount of the protein species in the composition inquestion.

The mode of separation is Size Exclusion Chromatography (SEC) and themethod uses 6M Guanidine HCl buffer as both sample solvent and HPLCmobile phase. Mercaptoethanol is used as a reducing agent to reduce thedisulphide (S—S) in the proteins or protein aggregates to createunfolded monomeric structures.

The sample preparation is easily achieved by dissolving 10 mg proteinequivalent in the mobile phase.

Two TSK-GEL G3000SWXL (7.7 mm×30.0 cm) columns (GPC columns) and a guardcolumn are placed in series to achieve adequate separation of the majorproteins in raw materials.

The eluted analytes are detected and quantified by UV detection (280nm).

Equipment/Materials:

-   1. HPLC Pump 515 with manual seal wash (Waters)-   2. HPLC Pump Controller Module II (Waters)-   3. Autosampler 717 (Waters)-   4. Dual Absorbance Detector 2487 (Waters)-   5. Computer software capable of generating quantitative reports    (Empower 3, Waters)-   6. Analytical column: Two TSK-GEL G3000SWXL (7.8×300 mm, P/N:    08541). Guard Column: TSK-Guard Column SWxL (6.0×40 mm, P/N: 08543).-   7. Ultrasonic Bath (Branson 5200)-   8. 25 mm Syringe filter with 0.2 μm Cellulose Acetate membrane.    (514-0060, VWR)

Procedure: Mobile Phase: A. Stock Buffer Solution.

-   -   1. Weigh 56.6 g of Na₂HPO₄, 3.5 g of NaH₂PO₄, and 2.9 g of EDTA        in to a 1000 mL beaker. Dissolve in 800 mL of water.    -   2. Measure pH and adjust to 7.5±0.1, if necessary, with HCl        (decrease pH) or NaOH (increase pH).    -   3. Transfer to a 1000 mL volumetric flask and dilute to volume        with water.

B. 6M Guanidine HCl Mobile Phase.

1. Weigh 1146 g of Guanidine HCl in to a 2000 mL beaker, and add 200 mLof the stock buffer solution (A)2. Dilute this solution to about 1600 mL with water while mixing with amagnetic stir bar (50° C.)3. Adjust the pH to 7.5±0.1 with NaOH.4. Transfer into a 2000 mL volumetric flask and dilute to volume withwater.5. Filter using the solvent filtration apparatus with the 0.22 μmmembrane filter.

Calibration Standards.

Calibration standards of each protein to be quantified are prepared thefollowing way:

-   -   1. Weigh accurately (to 0.01 mg) about 25 mg of the protein        reference standard into a 10 mL volumetric flask and dissolve in        10 mL of water.        -   This is the protein stock standard solution (S1) of the            protein    -   2. Pipette 200 μl of S1 into a 20 ml volumetric flask and dilute        to volume with mobile phase. This is the low working standard        solution WS1.    -   3. Pipette 500 μL of S1 into a 10 mL volumetric flask and dilute        to volume with mobile phase. This is standard solution WS2.    -   4. Pipette 500 μL of S1 into a 5 mL volumetric flask and dilute        to volume with mobile phase. This is standard solution WS3.    -   5. Pipette 750 μL of S1 into a 5 mL volumetric flask and dilute        to volume with mobile phase. This is standard solution WS4.    -   6. Pipette 1.0 mL of S1 into a 5 mL volumetric flask and dilute        to volume with mobile phase. This is the high working standard        solution WS5.    -   7. Using graduated disposable pipettes transfer 1.5 mL of WS1-5        into separate vials. Add 10 μL of 2-mercaptoethanol to each vial        and cap. Vortex the solutions for 10 sec. Let the standards stay        at ambient temperature for about 1 hr.    -   8. Filter the standards using 0.22 μm Cellulose Acetate syringe        filters.

The purity of protein is measured using Kjeldahl (N×6.38) and the area %from standard solution WS5 using the HPLC.

protein (mg)=“protein standard weight” (mg)×P1×P2

P1=P % (Kjeldahl)

P2=protein area % (HPLC)

Sample Preparation

-   -   1. Weigh the equivalent of 25 mg of protein of the original        sample into a 25 mL volumetric flask.    -   2. Add approximately 20 mL of mobile phase and let the sample        dissolve for about 30 min.    -   3. Add mobile phase to volume and add 167 μL of        2-mercaptoethanol to the 25 ml sample solution.    -   4. Sonicate for about 30 min and afterwards let the sample stay        at ambient temperature for about 1½ hours.    -   5. Mix the solution and filter using 0.22 μl Cellulose Acetate        syringe filters.

HPLC System/Columns Column Equilibration

-   1. Connect the GPC guard column and the two GPC analytical columns    in series.    -   New columns are generally shipped in a phosphate-salt buffer.-   2. Run water through a new column gradually from 0.1 to 0.5 mL/min    in 30 to 60 mins. Continue flushing for about 1 hour.-   3. Gradually decrease flow rate from 0.5 mL/min to 0.1 mL/min and    replace with mobile phase in the reservoir.-   4. Increase pump flow rate gradually from 0.1 to 0.5 mL/min in 30 to    60 mins to avoid pressure shock and leave at 0.5 mL/min.-   5. Inject ten samples to allow the column to be saturated and wait    for the peaks to elute.    -   This will aid in the conditioning of the column.    -   This step is done without the need of waiting for each injection        to be complete before injecting the next.-   6. Equilibrate with the mobile phase at least 1 hour.    Calculation of the Results Quantitative determination of the    contents of the proteins to be quantified, e.g. alpha-lactalbumin,    beta-lactoglobulin, and caseinomacropeptide, is performed by    comparing the peak areas obtained for the corresponding standard    proteins with those of the samples. The results are reported as g    specific protein/100 g of the original sample or weight percentage    of the specific protein relative to the weight of the original    sample.

Example 2: Crystallization of Beta-Lactoglobulin from a Crude WheyProtein Concentrate Protocol:

Lactose depleted UF retentate derived from sweet whey from a standardcheese production process and filtered through a 1.2 micron filter wasused as feed for the BLG crystallization process. The sweet whey feedwas conditioned on an ultrafiltration setup using a Koch HFK-328 typemembrane with a 46 mil spacer feed pressure of 1.5-3.0 bar, using a feedconcentration of 21% TS (total solids)±5 and polished water (waterfiltered by reverse osmosis to obtain a conductivity of at most 0.05mS/cm) as diafiltration medium. The temperature of the feed andretentate during ultrafiltration was approx. 12 degrees C. The pH wasthen adjusted by adding HCl to obtain a pH of approx. 5.40.Diafiltration continued until the drop in conductivity of the retentatewas below 0.03 mS/cm over a 20 min period. The retentate was thenconcentrated to approx. 30% TS (approx. 23.1% total protein relative tothe total weigh of the concentrated retentate). A sample of theconcentrated retentate was centrifuged at 3000 g for 5 minutes but novisible pellet was formed. The supernatant was subjected to HPLCanalysis. The composition of the feed is shown in Table 1.

The concentrated retentate was seeded with 0.5 g/L pure BLG crystalmaterial obtained from a spontaneous BLG crystallization (as describedin Example 3 in the context of feed 2). The seeding material wasprepared by washing a BLG crystal slurry 5 times in milliQ water,collecting the BLG crystals after each wash. After washing, the BLGcrystals were freeze dried, grounded up using a pestle and mortar, andthen passed through a 200 micron sieve. The crystallization seedstherefore had a particle size of less than 200 micron.

The concentrated retentate was transferred to a 300 L crystallizationtank where it was cooled to about 4 degrees C. and kept at thistemperature overnight with gentle stirring. Next morning, a sample ofthe cooled concentrated retentate was transferred to a test tube andinspected both visually and microscopically. Rapidly sedimentingcrystals had clearly formed overnight. A lab sample of the mixturecomprising both crystals and mother liquor was further cooled down to 0degrees C. in an ice water bath. The mother liquor and the crystals wereseparated by centrifugation at 3000 g for 5 minutes, and samples of thesupernatant and pellet were taken for HPLC analysis. The crystals werewashed once in cold polished water and then centrifuged again beforefreeze-drying the pellet.

TABLE 1 Concentration of selected components of the feed standardized to95% w/w total solids. Feed standardized to 95% TS Protein composition (%w/w relative to total protein)   ALA 17.7 BLG 51.6 CMP 19.5 Othercomponents (% w/w relative to total weight of the standardized feed) Ca0.357 K 0.200 Mg 0.058 Na 0.045 P 0.280 fat 5.6 protein 79

BLG Relative Yield Quantification by HPLC:

All samples were subjected to the same degree of dilution by addingpolished water. The samples were filtered through a 0.22 micron filter.For each sample, the same volume was loaded on an HPLC system with aPhenomenex Jupiter® 5 μm C4 300 Å, LC Column 250×4.6 mm (PartNumber:00G-4167-E0) and detected at 214 nm.

The samples were run using the following conditions:

Buffer A: MilliQ water, 0.1% w/w TFABuffer B: HPLC grade acetonitrile, 0.085% w/w TFAFlow: 1 ml/minGradient: 0-30 minutes 82-55% A and 18-45% B; 30-32 minutes 55-10% A and45-90% B; 32.5-37.5 minutes 10% A and 90% B; 38-48 minutes 10-82% A and90-18% B.

Data Treatment:

As all samples were treated in the same way, and we can directly comparethe area of the BLG peaks to gain a relative yield. As the crystals onlycontain BLG and the samples all have been treated in the same way, theconcentration of alpha-lactalbumine (ALA) and hence the area of ALAshould be the same in all of the samples. Therefore the area of ALAbefore and after crystallization is used as a correction factor (cf)when calculating the relative yield.

${cf_{\alpha}} = \frac{{area}\mspace{14mu}{of}\mspace{14mu}{ALA}_{{before}\mspace{20mu}{crystallization}}}{{area}\mspace{14mu}{of}\mspace{14mu}{ALA}_{{after}\mspace{14mu}{crystallization}}}$

The relative yield is calculated by the following equation:

${Yield}_{BLG} = {\left( {1 - \frac{cf_{\alpha} \times {area}\mspace{14mu}{of}\mspace{14mu}{BLG}_{{after}\mspace{14mu}{crystallization}}}{{area}\mspace{14mu}{of}\mspace{14mu}{BLG}_{{before}\mspace{20mu}{crystallization}}}} \right) \times 100}$

Results:

From the obtained chromatograms from before and after crystallization ofBLG from a sweet whey it is apparent that a large decrease in theconcentration of BLG has occurred, and using the yield calculation aspreviously described the yield of removed BLG was determined to 64.5%w/w.

The crystal slurry was investigated by microscopy. The sample containedhexagonal crystals, many having a size considerably larger than 200micron indicating that the observed crystals are not only the seedingcrystals. The crystals easily shattered when pressed with a needle whichconfirmed that they were protein crystals.

A chromatogram of a washed crystal product show that BLG makes up 98.9%of the total area of the chromatogram. The purity of the BLG product canbe increased even further by additional washing.

Conclusion:

This example demonstrates that surprisingly, it is possible tocrystalize BLG selectively from a crude whey protein concentrate whichcontains more that 48% non-BLG protein relative to total protein andthat the obtained BLG crystal isolate has an extremely high purity. Thisdiscovery opens up for a new approach for industrial milk proteinseparation, in which BLG is separated from the other protein componentsin a gentle way that preferably avoids extended exposure to hightemperatures and problematic chemicals.

Example 3: Crystallisation of BLG in Three Types of Whey ProteinSolutions Protocol:

Using the same experimental and analytical setup as in Example 2, threedifferent types of whey protein-containing raw material were tested asfeeds for crystallization. However, no seeding was used in theexperiment performed with feed 2. Feed 1 and 2 were based on sweet wheyand had been fat-reduced via a Synder FR membrane prior to treatment, asdescribed in Example 2. Feed 3 was derived from an acid whey.

The composition of the three feeds can be seen below in Table 2, Table3, and Table 4. Feed 3 was crystalized at 21% TS (total protein of 13.3%w/w relative to the total weight of the feed), a significantly lowerconcentration than the other two (total protein of 26.3% w/w in feed 1and 25.0% w/w in feed 2).

The slurry of the crystallized feed 1 was centrifuged on a Maxi-Spinfilter with a 0.45 micron CA membrane at 1500 g for 5 minutes. Then 2volumes of MilliQ water were added to the filter cake before it wascentrifuged again. The resulting filter cake was analyzed by HPLC. Thepellet from feed 2 was washed with 2 volumes of MilliQ water andcentrifuged again under standard conditions before the pellet wasanalyzed by HPLC. The pellet from feed 3 was analyzed without washing.

Crystals made from feed 2 were diluted to 10% TS and pH adjusted to pH 7using 1M NaOH to reverse the crystallization. NaCl was added to acrystal slurry from feed 2, 36% TS to reverse the crystallization.

TABLE 2 The concentration of selected components of feed 1 (whey proteinconcentrate based on sweet whey). Feed 1 (standardized to 95% TS)Protein composition (% w/w relative to total protein)   ALA 23.0 BLG55.1 CMP 20.5 Other components (% w/w relative to the total weight ofthe standardized feed) Ca 0.387 K 0.290 Mg 0.066 Na 0.068 P 0.207 FatBDL protein concentration 90 BDL = below detection limit in wet sample

TABLE 3 The concentration of selected components of feed 2 (ALA-reducedwhey protein concentrate based on sweet whey). Feed 2 standardized to95% TS Protein composition (% w/w relative to total protein)   ALA 12.2BLG 70.0 CMP 17.1 Other components (% w/w relative to the total weightof the standardized feed) Ca 0.387 K 0.204 Mg 0.066 Na 0.051 P 0.174 fatBDL protein concentration 89 BDL = below detection limit in wet,non-standardized sample.

TABLE 4 The concentration of selected components of feed 3 (whey proteinconcentrate based on acid whey). Feed 3 standardized to 95% TS Proteincomposition (% w/w relative to total protein) ALA 24.0 BLG 63.6 Otherwhey proteins 12.4 Other components (% w/w relative to the total weightof the standardized feed) Ca 0.205 K 0.051 Mg 0.013 Na 0.108 P 0.240 fat5.1 protein concentration 79

Results: Feed 1:

From chromatograms of the protein composition of the feed and the motherliquor it is evident that a large portion of BLG was recovered ascrystals by the process. The yield (calculated as described in example2) of isolated BLG is approx. 65% relative to the total amount of BLG inthe feed.

From a microscope photo of a sample taken during the early stages of thecrystallization period and a microscope photo of a sample which wastaken when the crystallization had ended, it is clear that the BLGcrystals are relatively fragile. Some of the crystals appear to breakduring stirring and are converted from hexagonal or rhombic shape tocrystals fragment which still appear very compact and well-defined buthave more irregular shapes.

From a chromatogram of the BLG crystals which was separated and washedon a spin filter it is seen that the purity is very high and the removalof other whey proteins is extremely efficient.

Feed 2:

From chromatograms of the protein composition of feed 2 and the obtainedmother liquor, it is evident that a large portion of BLG has beenremoved, and the calculated yield was 82% relative to the total amountof BLG in the feed 2.

By studying feed 2 before and after crystallization, it is seen thatduring crystallization the feed transformed from a transparent liquid(in which the stirring magnet was visible) to a milky white, opaqueliquid.

A microscope photo of the BLG crystals shows hexagonal shapes though themajority of the crystals are fractured.

A chromatogram of the isolated pellet of BLG crystals after being washedwith 2 volumes of MilliQ water clearly shows that the crystals containBLG in a very high purity.

Raising the conductivity (by adding NaCl) or altering the pH (byadjusting the pH to 7 by addition of NaOH) so that the environment nolonger favours the crystalline structure gives, in both cases show thatthe milky white suspension turns in to a transparent liquid as the BLGcrystals are dissolved.

The mineral composition of the crystal preparation obtained from feed 2is provided in Table 5. We note that the phosphorus to protein ratio wasvery low, which makes the crystal preparation suitable as a proteinsource for patients having kidney diseases.

TABLE 5 The concentration of selected components in the crystalpreparation obtained from feed 2. Composition of the crystal % w/wrelative to the preparation obtained from composition standardized feed2 to 95% TS Ca 0.119 K 0.047 Mg 0.019 Na BDL P BDL (less than 0.026)Total protein 93.4

Feed 3:

From chromatograms of the protein composition of feed 3 and theresulting mother liquor, it is evident that a large portion of BLG wasisolated (a calculated yield of 70.3% relative to the total amount ofBLG in the feed). If the protein content had been higher beforecrystallization, the obtained yield would have been even higher.

A microscope photo of the BLG crystals isolated from feed 3(substantially free of CMP) shows that the crystals had a rectangularshape as opposed to hexagonal. The rectangular crystals seemed morerobust than the hexagonal ones. For a chromatogram of the isolatedcrystal pellet without washing; the chromatogram clearly shows that thecrystals were BLG crystals despite having a rectangular shape instead ofa hexagonal shape

TABLE 6 The concentration of selected components of the crystalpreparation obtained from feed 3. Composition of the crystal % w/wrelative to the crystal preparation obtained preparation standardized tofrom feed 3 95% TS Ca 0.103 K BDL Mg 0.006 Na 0.035 P 0.041 Totalprotein 90

The crystal preparation derived from feed 3 contained 45 mg P/100 gprotein. We note that the phosphorus to protein ratio is very low, whichmakes the crystal preparation suitable as a protein source for patientshaving kidney diseases.

Conclusion:

All three feeds were suitable for the BLG crystallization process. TheBLG crystals were easily dissolved by adding salt or raising the pH orthe temperature. The new method makes it possible to prepare BLGpreparations with very low contents of phosphorus, which makes thepreparations suitable as a protein sources for patients having kidneydiseases.

Example 4: Preparation of Spray-Dried BLG Crystals and Determination ofBulk Density

A portion of the BLG crystals produced in Example 3 (using feed 2) wasseparated on a decanter centrifuge at 1200 g, 5180 RPM, 110 RPM Diff.with a 64 mil spacer (mil means 1/1000 inch) and a flow of 25-30 L/h.The BLG crystal phase was then mixed 1:1 with polished water and thenseparated again on the decanter centrifuge using the same settings. TheBLG crystal phase was then mixed with polished water in order to make itinto a slurry containing approx. 25% dry-matter and having acrystallinity of BLG of approx. 80, and subsequently dried on a pilotplant spray drier with an inlet temperature of 180 degrees C. and anexit temperature of 85 degrees C. without any preheating. Thetemperature of the liquid streams until spray-drying was 10-12 degreesC. The resulting powder sampled at the exit had a water content of 4.37%w/w.

The crystallinity of BLG in the slurry was approximately 90%.

The inventors have also successfully separated a slurry of BLG crystalsand mother liquor on a decanter centrifuge at 350 g, 2750 RPM, 150 RPMDiff. with a 64 mil spacer and a flow rate of 75 L/h. The BLG crystalphase was subsequently mixed 1:2 with polished water. The BLG crystalphase was then mixed with polished water in order to make it into athinner slurry, and subsequently dried on a pilot plant spray drierusing the same parameters as described above.

The bulk density of the spray-dried powder was then measured accordingto Example 1.17 and compared to the bulk density of a standard WPI driedon the same equipment. The standard WPI was found to have a bulk density(based on 625 stampings) of 0.39 g/mL, which is in the high end of thenormal range for a WPI powder. However, the spray-dried BLG crystalpreparation had a bulk density of 0.68 g/mL, more than 75% higher thanthe bulk density of the standard WPI. This is truly surprising andprovides a number of both logistic and application-related advantages.

TABLE 7 The concentration of selected components of the spray- dried BLGcrystal preparation of Example 7. BDL = below detection limit Spraydried BLG crystal powder Protein composition (% w/w relative to totalprotein) ALA 0.7 BLG 97.4 CMP BDL Other components (% w/w relative tototal weight of the BLG crystal powder) Ca 0.118 K 0.026 Mg 0.017 Na BDLP BDL water 3.8 protein concentration 94

A sample of the spray-dried BLG crystal preparation was subsequentlyresuspended in cold demineralised water and BLG crystals were stillclearly visible by microscopy. Addition of citric acid or NaCl causedthe BLG crystals to dissolve and transformed the opaque crystalsuspension into a clear liquid.

The inventors have seen indications that extended heating during thedrying step reduces the amount of BLG that is in crystal form. It istherefore preferred that the heat exposure of the BLG crystalpreparation is as low as possible.

Conclusion:

This example demonstrates that slurries comprising BLG crystals can bespray-dried and that BLG crystals are still present in the resuspendedspray-dried powder if the heating during the drying step is controlled.

The inventors furthermore found that the bulk density of a whey proteinpowder that contains BLG crystals is considerably higher than thatobtained by normal spray-drying of dissolved protein streams. Highdensity powders allows for more cost-effective packaging and logisticsof the powder as less packaging material is required per kg powder andmore powder (mass) can be transported by a given container or truck.

The high density powder also appears to be easier to handle and lessfluffy and dusty during manufacture and use.

Example 5: Low Phosphorus Protein Beverage

Six low phosphorus instant beverage powders were prepared using thepurified BLG product from Example 3 (the crystal preparation obtainedfrom feed 3). All the dry ingredients were blended to obtain an instantbeverage powder and then mixed with demineralized water to obtain 10 kgof each sample and allowed to hydrate for 1 hour at 10 degrees C.

TABLE 8 Composition of the six beverage samples. Beverage sampleIngredient % w/w A B C D E F Dried, purified 5.0 10.0 5.0 10.0 5.0 10.0BLG from Ex. 3, feed 3 Citric acid To To To To To To pH pH pH pH pH pH3.5 3.5 3.0 3.0 4.0 4.0 Sucrose 10.0 10.0 10.0 10.0 10 10 DemineralisedTo To To To To To water 100% 100% 100% 100% 100% 100%

The turbidity of the sub-samples of the six samples was measured on aTurbiquant® 3000 IR Turbidimeter and the viscosity on a vicoman byGilson. The results are shown in the table below.

TABLE 9 Measured viscosity and turbidity of the six beverage samples.Sample viscosity (Cp) NTU A 1.42 36.2 B 2.37 46.3 C 2.69 4.9 D 2.70 5.0E 1.45 63.1 F 2.25 82.1

A photo of test tubes containing sub-samples of the six low phosphorousbeverage samples is shown in FIG. 3. From left to right, the sub-sampleswere sample A, B, C, D, E, and F. The visual inspection of the testtubes verified the turbidity measurements and documented that allbeverage samples were transparent and that particularly samples C and D(pH 3.0) were very clear. The low viscosities demonstrate that thebeverage samples were easily drinkable.

All ingredients used for preparing the beverage were low in phosphorusand did not contain unnecessary minerals. The obtained beveragestherefore had a phosphorus content of approx. 45 mg P/100 g protein andgenerally had a very low mineral content. The six liquid food productsprepared from the instant beverage powder were therefore suitable foruse as instant protein beverages for kidney disease patients.

Example 6: Crystal Separation by Dynamic Cross-Flow Filtration

Lactose-depleted UF retentate derived from sweet whey from a standardcheese production process, filtered through a 1.2 micron filter, wasused as feed for the crystallization process. The sweet whey feed wasconditioned on an ultrafiltration setup using a Koch HFK-328 typemembrane with a 46 mil spacer, a feed pressure of 1.5-3.0 bar, using afeed concentration of 10% TS (total solids)±5, and polished water (waterfiltered by reverse osmosis to obtain a conductivity of at most 0.05mS/cm) as diafiltration medium. The temperature of the feed andretentate during ultrafiltration was approx. 12 degrees C. The pH wasthen adjusted by adding HCl to obtain a pH of approx. 5.60.Diafiltration continued until the conductivity of the retentate wasbelow 1.30 mS/cm. The feed was then heated to 25 degrees C. before theretentate was concentrated to approx. 27% TS (approx. 21% total proteinrelative to the total weigh of the concentrated retentate). The permeateconductivity was 0.33 mS/cm at the end of the concentration. A sample ofthe concentrated retentate was centrifuged at 3000 g for 5 minutes butno visible pellet was formed.

The concentrated retentate was transferred to a 300L crystallizationtank where it was cooled to about 6 degrees C. and kept at thistemperature overnight with gentle stirring. The next morning, theretentate had crystallized. The mother liquor and the crystals wereseparated by centrifugation at 3000 g for 5 minutes, and samples of thesupernatant and pellet were taken for HPLC analysis. The yield of BLGfrom this process was calculated to be 67%.

The crystal slurry from the 300 L tank was used for a feed in an AndritzDCF 152S system using one disk membrane with a pore size of 500 nm. Thefiltration was run at 8 degrees C., rotational speed was 32 Hz, and thetransmembrane pressure was 0.4 bar. The system works as a dead endfiltration where retentate is built up in the filtration chamber, unlikea larger unit where the retentate would be continuously removed. Thefiltration was run in a stable manner for just over 40 minutes, at whichpoint the solids, which had built up in the filtration chamber, startedto influence the filtration.

The amount of crystal mass increased significantly during the DFCoperation.

Conclusion: The DCF provides a stable and efficient means for separatingthe crystals from the ML. If needed, washing liquid could be added tothe DCF.

Example 7: Degree of Protein Denaturation of Different Whey ProteinProducts

The degree of protein denaturation of a commercial product and four BLGisolates were compared. The BLG isolates are suitable for preparing theinstant beverage powder of the invention. The samples are describedbelow.

  Samples A: BiPro (Commercially available WPI; Davisco, USA) B: BLGcrystal slurry as is - no drying (invention) C: BLG crystal slurryfreeze dried (invention) D: BLG crystals redissolved (pH 7) andfreeze-dried E: BLG crystal slurry spray dried (invention)

Samples B-E were prepared the following way:

Crystal slurry was prepared as described in Example 6 and separated asdescribed in Example 4. Some of the separated BLG slurry was taken outand split into four portions.

Sample B: The first portion of the separated BLG crystal slurry wasre-dissolved without any drying by adjusting the pH of the BLG crystalslurry to 7.01 using a 3% NaOH; and the sample was then diluted to Brix6 in order to make an approximately 5% protein solution.

Sample C: The second portion of the separated BLG crystal slurry wasfreeze-dried. The powder was then re-suspended in polished water, the pHwas adjusted to 7.09 using a 3% NaOH, and the sample was then diluted toBrix 6 in order to make an approximately 5% protein solution.

Sample D: The third portion of the separated BLG crystal slurry wasre-dissolved by adjusting the pH to 7.0 using a 3% NaOH, and thenfreeze-dried. The freeze-dried powder was then resuspended in polishedwater, and the pH was measured to be 7.07. The sample was then dilutedto Brix 6 in order to make an approximately 5% protein solution.

Sample E: The fourth portion of the separated BLG crystal slurry wastreated and spray dried as described in Example 4. The powder was thenre-suspended in polished water, and the pH was adjusted to 7.04 using a3% NaOH. The sample was then diluted to Brix 6 in order to make anapproximately 5% protein solution.

The degree of protein denaturation of each sample was determinedaccording to Example 1.3 and the results are presented in the tablebelow.

TABLE Comparing the degree of protein denaturation of a commerciallyavailable WPI product (Bipro) with 4 BLG products which can be used inthe instant beverage powder of the invention. Total Total con- proteincentration Degree concen- of soluble of protein tration proteindenaturation Sample at pH 7 at pH 4.6 (%) A: BiPro (Commerciallyavailable WPI) 5.11 4.54 11.15 B: BLG crystal slurry as is (no drying)4.62 4.56 1.30 C: BLG crystal slurry freeze-dried 4.74 4.69 1.05 D: BLGcrystals re-dissolved (pH 7) 4.74 4.69 1.05 and freeze-dried E: BLGcrystal slurry spray-dried 4.75 4.71 0.84

Conclusion:

Regardless of the drying method, the BLG isolate have a surprisingly lowdegree of denatured protein; only a tenth of what can be found in thecommercially available WPI used for comparison. It is particularlysurprising that the spray-dried BLG crystal slurry product still has thelowest degree of denaturation of all products.

Example 8: Production of a Spray-Dried, Acidic BLG Isolate Powder WheyProtein Feed

Lactose-depleted UF retentate derived from sweet whey from a standardcheese production process was filtered through a 1.2 micron filter andhad been fat-reduced via a Synder FR membrane prior to being used asfeed for the BLG crystallisation process. The chemical composition ofthe feed can be seen in Table 10. We note that all weight percentages ofspecific proteins, such as BLG, ALA, mentioned in this Example pertainto the weight percentage of the non-aggregated proteins relative tototal protein.

Conditioning

The sweet whey feed was conditioned on an ultrafiltration setup at 20degrees C., using a Koch HFK-328 type membrane (70 m² membrane) with a46 mill spacer feed pressure 1.5-3.0 bar, to a feed concentration of 21%total solids (TS)±5, and using as diafiltration medium polished water(water filtered by reverse osmosis to obtain a conductivity of at most0.05 mS/cm). The pH was then adjusted by adding HCl so that the pH wasapprox. 5.5. Diafiltration continued until the drop in conductivity ofthe retentate was below 0.1 mS/cm over a 20 min period. The retentatewas then concentrated until the permeate flow was below 1.43 L/h/m². Afirst sample of concentrated retentate was taken and subjected tocentrifugation at 3000 g for 5 minutes. The supernatant of the firstsample was used for the determination of BLG yield.

Crystallisation

The concentrated retentate was transferred to a 300 L crystallisationtank where it was seeded with pure BLG crystal material made fromrehydrated, spray-dried BLG crystals. Subsequently, the seeded wheyprotein solution was cooled from 20 degrees C. to approx. 6 degrees C.over approx. 10 hours to allow the BLG crystals to form and grow.

After cooling, a sample of the crystal-containing whey protein solution(the second sample) was taken and the BLG crystals were separated bycentrifugation at 3000 g for 5 minutes. The supernatant and crystalpellets from the second sample were subjected to HPLC analysis asdescribed below. The yield of crystallization was calculated as outlinedbelow and determined to 57%.

TABLE 10 Chemical composition of the feed Feed standardized to 95% totalsolids Protein composition % w/w of total protein ALA 10.2 BLG 59.6Other proteins 30.2 Selected other components % w/w Ca 0.438 K 0.537 Mg0.077 Na 0.131 P 0.200 Fat 0.220 protein concentration 87

BLG Yield Determination Using HPLC:

The supernatants of the first and second samples were subjected to thesame degree of dilution by adding polished water and the dilutedsupernatants were filtered through a 0.22 μm filter. For each filteredand diluted supernatant the same volume was loaded on an HPLC systemwith a Phenomenex Jupiter® 5 μm C4 300 Å, LC Column 250×4.6 mm, Ea. anddetected at 214 nm.

The samples were run using the following conditions:

Buffer A: MilliQ water, 0.1% w/w TFABuffer B: HPLC grade acetonitrile, 0.085% w/w TFAFlow: 1 mL/minColumn temperature: 40 degrees C.Gradient: 0-30 minutes 82-55% A and 18-45% B; 30-32 minutes 55-10% A and45-90% B; 32.5-37.5 minutes 10% A and 90% B; 38-48 minutes 10-82% A and90-18% B.

Data Treatment:

As both supernatants were treated in the same way, one can directlycompare the area of the BLG peaks to calculate a relative yield. As thecrystals only contain BLG and the samples all have been treated in thesame way, the concentration of alpha-lactalbumin (ALA) and hence thearea of ALA should be the same in all of the samples. Therefore, thearea of ALA before and after crystallisation is used as a correctionfactor (cf) when calculating the relative yield.

${cf_{\alpha}} = \frac{{area}\mspace{14mu}{of}\mspace{14mu}{ALA}_{{before}\mspace{20mu}{crystallization}}}{{area}\mspace{14mu}{of}\mspace{14mu}{ALA}_{{after}\mspace{14mu}{crystallization}}}$

The relative yield is calculated by the following equation:

${Yield}_{BLG} = {\left( {1 - \frac{cf_{\alpha} \times {area}\mspace{14mu}{of}\mspace{14mu}{BLG}_{{after}\mspace{14mu}{crystallization}}}{{area}\mspace{14mu}{of}\mspace{14mu}{BLG}_{{before}\mspace{20mu}{crystallization}}}} \right) \times 100}$

Acid Dissolution of BLC Crystals

The remainder of the material from the crystallisation tank wasseparated using a decanter at 350 g, 2750 RPM, 150 RPM Diff. with a 64spacer and a feed flow of 75 L/h before separation the feed was mixed1:2 with polished water. The BLG crystal/solid phase from the decanterwas then mixed with polished water in order to make it into a thinnerslurry before a phosphoric acid was added to lower the pH to approx. 3.0in order to quickly dissolve the crystals.

After dissolving the BLG crystals, the pure BLG protein liquid wasconcentrated to 15 Brix on the same UF setup as used to prepare the feedfor crystallisation and the pH was adjusted to final pH of approx. 3.8.The liquid BLG isolate was then heated to 75 degrees for 5 minutes andsubsequently cooled to 10 degrees C. The heat-treatment was found toreduce the microbial load from 137.000 CFU/g prior to the heat-treatmentto <1000 CFU/g after the heat-treatment. The heat-treatment did notcause any protein denaturation and the intrinsic tryptophan fluorescenceratio (330 nm/350 nm) was determined to 1.20 indicating nativeconfirmation of the BLG molecules.

The BLG was dried on a pilot plant spray drier with an inlet temperatureof 180 degrees C. and an exit temperature of 75 degrees C. The resultingpowder sampled at the exit had a water content of approx. 4% w/w, thechemical composition of the powder is shown in Table 11. A sample of thedried powder was dissolved and the degree of protein denaturation wasdetermined to 1.5% and the intrinsic tryptophan fluorescence emissionratio (I330/I350) was measured to 1.20.

TABLE 11 The composition of the BLG isolate powder (BDL = below thedetection limit) BLG isolate powder standardized to 95% total solidsProtein composition % w/w of total protein ALA 0.4 BLG 98.2 Otherprotein 1.4 Other selected components % w/w Ca BDL K BDL Mg BDL Na BDL P0.781 fat 0.09 protein concentration 90

The bulk density (625 taps) of the spray-dried powder was estimated at0.2-0.3 g/cm³.

Conclusion: By using the above described process, we were able toproduce a high-purity BLG product that can be heat-treated withsubstantially no protein denaturation or protein unfolding duringprocessing. The heat-treatment greatly lowered the bacteria levelswithout damaging the protein product.

The inventors have seen indications that even higher bulk density can beobtained by increasing the protein content prior to spray-drying. Also,the inventors have observed that even lower degrees of denaturation areobtained if the entry and/or exit temperature used for spray-drying arereduced.

Example 9: Production of a Spray-Dried, pH-Neutral BLG Isolate Powder

When using the same protocol and experimental setup as in Example 2, thelactose-reduced whey protein isolate shown in Table 12 was conditionedand used for feed for crystallization. The yield of crystallization wascalculated to be 68%.

We note that all weight percentages of specific proteins, such as BLGand ALA, mentioned in this Example pertain to the weight percentage ofthe non-aggregated proteins relative to total protein.

TABLE 12 Composition of the feed FEED standardized to 95% total solidsProtein composition % w/w of total protein ALA 9.1 BLG 59.1 Otherprotein incl. CMP 31.6 Other selected components % w/w Ca 0.445 K 0.574Mg 0.074 Na 0.128 P 0.211 fat 0.513 protein concentration 84

The remainder of the material from the crystallization tank wasseparated on a decanter at 350 g, 2750 RPM, 150 RPM Diff. with a 64spacer and a feed flow of 75 L/h. before separation the feed was mixed1:2 with polished water. The BLG crystal/solid phase from the decanterwas then mixed with polished water in order to make it into a thinnerslurry before 0.1 M potassium hydroxide was added to adjust the pH toapprox. 7 in order to quickly dissolve the crystals.

After dissolving the crystals, the pure BLG protein liquid wasconcentrated to brix 15 on a the same UF setup as used to prepare thewhey protein solution for crystallization and the pH was adjusted to thefinal pH of 7.0. The BLG was dried on a pilot plant spray drier with aninlet temperature of 180 degrees C. and an exit temperature of 75degrees C. The resulting powder sampled at the exit had a water contentof approx. 4% w/w. The composition of the powder is shown in Table 13.After drying, some of the powder was dissolved in demineralized waterand the degree of protein denaturation was determined to 9.0% and theintrinsic tryptophan fluorescence ratio (330 nm/350 nm) was 1.16.

TABLE 13 Chemical composition of the BLG isolate powder BLG isolatepowder standardized to 95% total solids Protein composition % w/w oftotal protein ALA 0.2 BLG 98.9 Other protein 0.9 Other selectedcomponents (% w/w) Ca 0.003 K 2.343 Mg BDL Na BDL P 0.629 fat 0.329protein concentration 88

The bulk density (625 taps) of the spray-dried powder was estimated at0.2-0.3 g/cm³.

Conclusion: By using the above described process, we are able to producea pH-neutral, high-purity BLG product with minimum to no proteindenaturation during processing. The inventors have seen indications thateven higher bulk density can be obtained by increasing the proteincontent prior to spray-drying. Also, the inventors have observed thateven lower degrees of denaturation are obtained if the entry and/or exittemperature used for spray-drying are reduced. The level of denaturationmay furthermore be reduced by reducing the mineral content prior tospray-drying.

Example 10: Preparation of Coated BLG Isolate Powder

Using a spray-dried BLG isolate, produced as described in example 4 orexample 9, a coated BLG isolate was produced in a fluid bed (DIOSNA,MINILAB XP no. 365-1461). The inlet temperature was 60 degrees C. Thepowder temperature was between 40 and 50 degrees C. for the duration ofthe process and the air flow was 25-35 m³ per hour. The coating materialwas dissolved in 50 g of demineralized water and slowly injected intothe fluid bed, where it was nebulized. For each batch, 500 g of thespray dried BLG isolate was used. After the coating material had beenadded, the drying continued until the coated BLG isolate had a moisturecontent of 4-5%. Using this setup a BLG isolate coated with 25 g ofcitric acid and a BLG isolate coated with 30 g of trisodium citrate wasproduced.

Test samples were prepared as shown in the table and analyzed withregard to solubility as described below. In addition to the 10% w/wsolution described, a 30% w/w solution of BLG isolate powder wereprepared and tested. The test samples were further analyzed with regardto wettability as described below. In addition the test samples wereevaluated sensorically by two trained test persons according to theparameters set out in example 1.11.

Using the BLG isolate powder from example 8, a powder coated withlecithin was produced in the same fluid bed as above. 500 g of powderwas added to the fluid bed inlet temperature was 75 degrees C. and theair flow was 25 m³/h. When the powder temperature reached 38 degrees C.,50 mL of water was added slowly via the nebulizer. The powdertemperature was allowed to rise up to 45 degrees C., and 5 ml lecithinwas injected through the nebulizer. The powder was heated to 65 degreesC. and dried until it contained less than 5% moisture.

Solubility test: The solubility and readiness of an instant powder todissolve can be measured by the present test. 10 gram of the powder isadded to 90 grams of demineralized water (8 degrees C.) in a sealabletransparent test tube. The mixture is shaken vigorously by hand for 30seconds. The mixture is evaluated immediately and left to stand for 1minute, whereafter the mixture is evaluated again. The evaluation iscarried out by visual inspection of the following parameters:transparency of liquid phase, foam formation, color and to which extendthe powder has dissolved.

Results

Sample # Powder % w/w 1 10% neutral uncoated BLG isolate, pH 7.02 2 10%WPI coated with lecithin 3 10% BLG isolate coated with lecithin 4 10%BLG isolate coated with Citric acid 5 10% BLG isolate coated withtrisodium citrate 6 10% standard WPI, no coating 7 30% BLG coated w.trisodium citrate 8 30% neutral BLG isolate, no coating, pH 7.02 9 30%standard WPI, no coating

After 1 minutes # First evaluation evaluation Taste 1 Easily dissolved,Stable foam, Whey taste: 0-1, Foam formation, volume increase in novisible particles liquid phase, liquid in the liquid or in phasecompletely the foam clear 2 Easily dissolved, Stable foam but less Wheytaste: 7 with Foam formation, foam compared to some undertaste novisible particles the other test mixtures. in the liquid or in liquidphase turbid the foam, some and a bit yellow yellow color 3 Easilydissolved, Stable foam but less whey taste: 1-2, with Foam formation,foam compared to some undertaste, as #2 no visible particles sample 2.liquid phase in the liquid or in turbid and a bit the foam, slightlyyellow yellow color 4 Easily dissolved, Stable foam, pleasant Foamformation, liquid phase turbid Whey taste: 0-1 no visible particles butno discoloration Citric acid: 6 in the liquid or the foam 5 Easilydissolved, Stable foam, Slight taste of Foam formation, liquid phaseslightly minerals no visible particles turbid but no Whey taste:2-3 inthe liquid or the discoloration foam 6 Easily dissolved, Stable foam,whey taste: 8 Foam formation, slightly yellow color no visible particlesin the liquid or the foam, slightly yellow color 7 Easily dissolved,Stable foam, Not tasted Foam formation, liquid phase turbid no visibleparticles but no discoloration in the liquid or the foam 8 Easilydissolved, Stable foam, Neutral but with a Foam formation, Volumeincrease in milky hint and a bit no visible particles liquid phase,liquid protein taste in the liquid or the phase completely Whey taste: 6foam clear 9 Foam formation, Stable foam, Not tasted some small visiblesome yellow particles in the discoloration foam, some yellowdiscoloration

Conclusion: The BLG isolate powders procured had a good agglomeratedstructure and easily dissolved in water. All coated BLG isolate powders(samples 1, 3-5 and 7-8) performed equal to or better than a normallycoated WPI (test sample 2) in this test. When testing the 30% version itwas surprisingly easy to dissolve the BLG isolates, both coated anduncoated (test samples 1, 3-5 and 7-8) and therefore it is believed thatthere is a possibility to go even higher than 30% in proteinconcentration. Both the coated and uncoated BLG isolates in 30% solution(samples 7-8) were easily dissolved as compared with the standard WPI(sample 9), which had visible particles in the foam.

Wettability: The method is used to describe the wettability of a powder.The wettability is defined as the time it takes before the entire sampleis wet. 0.5 grams of the powder is measured out and placed on thesurface of 100 g of demineralized water (5 degrees C.) in a cylindricalcontainer with a diameter of 5 cm. The time from placing the powder onthe surface of the water to the powder was dissolved or has passedthrough the water surface is measured.

Results

Time to Sample dissolve # Powder (minutes) comments 10 Neutral BLG 22The powder was not as evenly isolate, uncoated distributed over thesurface and this is believed to be the reason for the slightly slowerwetting 11 WPI coated with. 18 lecithin 12 Neutral BLG 16 isolate coatedwith lecithin 13 BLG isolate +80 Almost no wetting occurred. coated withCitric acid 14 BLG isolate 17 coated with. trisodium citrate 15 StandardWPI +80 After 80 minutes the test was terminated and roughly 2/3 of theWPI had been dissolved at this point

Conclusion: The coated BLG isolate powders (sample 10, 12, 14) wetted ina similar way as at normal instantized WPI (samples 11, 15) with theexception of the BLG isolate powder coated with citric acid (sample 13).It was surprising that non coated BLG isolate powder wetted much betterthan a standard WPI (sample 6).

Example 11: Wettability of Uncoated BLG Isolates

In the present example, the wettability of an uncoated acidic BLGisolate and an uncoated neutral BLG isolate is compared with thewettability of an uncoated whey protein isolate (WPI), The wettabilityis defined as the time it takes before the entire sample is wet. 0.5grams of the powder is measured out and placed on the surface of 100 gdemineralized water (10 degrees C.) in a cylindrical container with adiameter of 5 cm. The time from placing the powder on the surface of thewater to the powder was dissolved or has passed through the watersurface is measured.

Results

Sample Time to dissolve # Powder (minutes) comments 1 WPI +55 5-10% ofthe powder was still left on the surface after 55 minutes 2 acidic BLGisolate 27 Completely wetted and (pH3.7), uncoated dissolved 3 neutralBLG 15 Completely wetted and isolate(pH7), uncoated dissolved

Conclusion: It was surprising that uncoated BLG isolate powder (samples2, 3) wetted much better than a standard WPI (sample 1).

Example 12: Preparation of Instant Beverage Powder

Preparation of an Instant Powder Used as a Nutritional Supplement.

100 g of the instant powder is prepared by blending the followingingredients: 91 grams of whey protein concentrate comprising at least85% w/w of BLG as prepared in example 4, and 4 grams of soja lecithin.

100 grams of the instant powder contains 360 Kcal with an energydistribution as follows: 2 E % from lipid, 1 E % from carbohydrate and97 E % from protein. Food products prepared from the instant powder canbe used as a protein supplement for treatment of patients with or atrisk of malnutrition e.g. by mixing the instant powder with water toobtain a beverage or by adding the powder to regular meals. 10-15 gramsof the instant powder is stirred into water having a temperature of15-25 degrees C. The prepared instant powder drink has a pleasant taste,colour and viscosity.

Example 13: Preparation of Instant Beverage Powder Preparation of aNutritionally Complete Instant Powder Comprising BLG.

100 g of the instant powder is prepared by blending 18.2 grams ofvegetable oil (a mixture consisting of palm kernel oil, coconut oil,rapeseed oil and sunflower oil), 56.4 grams of glucose syrup, and 20grams of whey protein concentrate comprising at least 90% w/w of BLG asprepared in example 4 and about 3 gram soja lecithin.

Further is added: potassium citrate, sodium citrate, sodium chloride,magnesium hydrogen phosphate, potassium hydrogen phosphate, magnesiumchloride, cholin chloride, calcium carbonate, calcium phosphate, sodiumL-ascorbate, aroma, iron sulphate, 1-ascorbin acid, zinc sulphate,magnesium citrate, DL-alpha-tocopheryl acetate, manganese sulphate,nicotin amide, Dbiotin, copper sulphate, calcium-D-pantothenate, pteroylmonoglutaminic acid, sodium fluoride, DL-alpha-tocopherol, thiaminhydrochloride, pyridoxine hydrochloride, carotenoids, retinyl palmitate,riboflavin, cyanocobalamin, cholecalciferole, chrom chloride, sodiummolybdenum, potassium iodide, sodium selenite and phytomenadion toobtain an instant powder having the following nutritional profile: 381μg RE vitamin A, 0.91 mg carotenoids, 3.3 μg vitamin D, 5.9 mg α-TEvitamin E, 24 μg vitamin K, 0.69 mg vitamin B1, 0.74 mg vitamin B2, 8.3mg-NE niacin, 2.5 mg pantothenic acid, 0.79 μg vitamin B6, 123 μg folicacid, 0.98 μg vitamin B12, 24 μg biotine, 60 mg vitamin C, 156 mgcholin, 1.17 gram salt, 469 mg sodium, 705 mg potassium, 578 mgchloride, 371 mg calcium, 337 mg phosphorous, 107 mg magnesium, 7.4 mgiron, 5.6 mg zinc, 0.83 mg copper, 1.5 mg manganese, 0.5 g fluoride, 48μg molybdenum, 26 μg selenium, 25 μg chrome, 61 μg iodide.

The instant powder contains 462 Kcal/100 gram with an energydistribution as follows: 35.6 E % from lipid, 48.7 E % from carbohydrateand 15.7 E % from protein. Food products prepared from the instantpowder can be used as nutritionally complete food products for treatmentof patients with or at risk of malnutrition e.g. by mixing the instantpowder with water to obtain a drink or to be used for tube feeding. 22gram of the instant powder is stirred into 85 ml cold water (which hasbeen boiled) to obtain a drink containing 100 kcal. 33 grams of theinstant powder is stirred into 78 ml cold water (which has been boiled)to obtain a drink containing 150 kcal. The prepared instant powderbeverage has a pleasant taste, colour and viscosity.

1. An instant beverage powder comprising at least 1% w/wbeta-lactoglobulin (BLG), preferably at least 5%, wherein: i. thecrystallinity of BLG is at least 20%, preferably at least 40%, and/orii. at least 85% w/w of the total amount of protein is comprised by BLG,and furthermore comprising at least one additional ingredient selectedfrom the group consisting of vitamins, flavouring agent, colouringagent, minerals, sweeteners, antioxidants, food acid, lipids,carbohydrate, prebiotics, probiotics, anti-foaming agents and non-wheyprotein.
 2. The powder according to claim 1, wherein the instantbeverage powder comprises BLG in the range of 1-90% % w/w BLG.
 3. Thepowder according to any of claims 1-2, wherein said powder furthercomprises one or more of: i. a sweetener, e.g. a sugar sweetener and/ora non-sugar sweetener, ii. a flavoring agent, iii. at least one foodacid, e.g. citric acid or other suitable food acids, iv. the sum of theamounts of Na, K, Mg, and Ca of the instant beverage is at most 10mmol/g protein and wherein a 10% w/w solution of the powder indemineralized water has a pH in the range of 2-8.
 4. The powderaccording to any of the preceding claims, wherein said powder comprisesat least 85% w/w BLG relative to total protein such as at least 86% w/wBLG relative to total protein, at least 87% w/w BLG relative to totalprotein, at least 88% w/w BLG relative to total protein, at least 89%w/w BLG relative to total protein.
 5. The powder according to any of thepreceding claims, wherein said powder comprises at least 91% w/w BLGrelative to total protein such as at least 92% w/w BLG relative to totalprotein, at least 93% w/w BLG relative to total protein, at least 94%w/w BLG relative to total protein, at least 95% w/w BLG relative tototal protein, at least 96% w/w BLG relative to total protein, at least97% w/w BLG relative to total protein, at least 98% w/w BLG relative tototal protein or at least 99% w/w BLG relative to total protein.
 6. Thepowder according to any of the preceding claims wherein the powdercomprises a water content in an amount of at most 6% w/w, such as atmost 5% w/w, preferably at most 4% w/w, more preferably at most 3% w/w,and even more preferably at most 2% w/w.
 7. The powder according to anyof the preceding claims, wherein said powder comprises: i. at most 6%w/w water ii. at least 15% w/w total protein relative to total solidsiii. at least 85% w/w BLG relative to total protein, wherein said powderis a dry powder.
 8. The powder according to any of the preceding claims,wherein said powder has a bulk density of at least 0.30 g/mL, preferablyat least 0.4 g/mL, more preferably 0.5 g/mL or even more preferably 0.5g/mL.
 9. The powder according to any of the preceding claims, whereinthe sum of ALA and caseinomacropeptide (CMP) comprises at least 40% w/wof the non-BLG protein of the powder, preferably at least 60% w/w, evenmore preferably at least 70% w/w, and most preferably at least 85% w/wof the non-BLG protein of the powder.
 10. The powder according to any ofthe preceding claims, wherein the sum of the amounts of Na, K, Mg, andCa is at most 10 mmol/g protein
 11. The powder according to any of thepreceding claims, wherein said powder has an energy content in the rangeof about 200-500 kcal/100 grams of powder.
 12. The powder according toany of the preceding claims, wherein the energy amount of the protein isat least 7 E %, preferably at least 25 E %, more preferably at least 30E %, even more preferably at least 40 E %.
 13. The powder according toany of the preceding claims, wherein the contribution of energy fromlipid is in the range of 0-60 E %.
 14. The powder according to any ofthe preceding claims, wherein the contribution of energy fromcarbohydrate is in the range of 0-90 E %.
 15. The powder according toany of the preceding claims, wherein said powder in a 10% w/w solutionin demineralised water has a pH in the range of 2-8 at room temperature.16. The powder according to any of the preceding claims, for use as afood ingredient.
 17. The powder according to any of the precedingclaims, for use in a method for the treatment of patients with or atrisk of malnutrition.
 18. The powder according to any of the precedingclaims, for use in a method for the treatment of kidney disease.
 19. Aliquid food product comprising a liquid and the instant beverage powderaccording to any of the claims 1-18.
 20. The food product according toclaim 19, wherein the product comprises at most 40 gram of said powderper 100 grams of said liquid, preferably at most 30 gram of said powderper 100 grams of said liquid.
 21. The food product according any ofclaims 19-20 having a turbidity of at most 200 NTU.
 22. The food productaccording any of claims 19-20 having a turbidity of more than 200 NTU.23. The food product according to any of claims 19-22, wherein saidliquid is selected from the group consisting of water, milk products,fruit juice, vegetable juice, beverages and combinations thereof. 24.The food product according to any of claims 19-23, wherein said foodproduct comprises water and the instant beverage powder according any ofclaims 1-18, said food product having an energy content in the range of30-300 kcal/100 grams of food product.
 25. The food product according toclaim 24, wherein said food product has an energy content in the rangeof 30-100 kcal/100 grams of food product, preferably in the range of30-100 kcal/100 grams of food product, more preferably in the range of40-90 kcal/100 grams of food product, or even more preferably in therange of 40-70 kcal/100 grams of food product.
 26. The food productaccording to claim 24, wherein said food product has energy content inthe range of 100-300 kcal/100 grams of food product, preferably in therange of 100-250 kcal/100 grams of food product, or in the range of125-225 kcal/100 grams of food product.
 27. The food product accordingto any of claims 19-26, wherein the protein fraction of the liquid foodproduct has a color value delta b* in the range of −0.10 to +0.51 at theCIELAB color scale, wherein deltab*=b_(sample standardized to 6.0 w/w % protein)−b_(demin. water),measured at room temperature.
 28. The food product according to any ofclaims 19-27, wherein the product further comprises vegetables and/orfruit.
 29. The food product according to any of claims 19-28, for use asa nutritional supplement.
 30. The food product according to any ofclaims 19-29, wherein said food is ingested before, during or afterexercise.
 31. The food product according to any of claims 19-30, for usein the treatment of patients with or at risk of malnutrition.
 32. Thefood product according to any of claims 19-31, for use in the treatmentof kidney disease.
 33. A kit comprising the powder according to any ofclaims 1-18, said kit comprising i. a tool for measuring said powder,and ii. a container having a lid for opening and closing the container,wherein said container is for mixing said powder with a liquid to form afood product, and said container is adapted for drinking the foodproduct directly from the container.
 34. A method for preparing a liquidfood product according to any of the claims 19-32, said methodcomprising i. Adding a powder according to any of claims 1-18 to aliquid, ii. Optionally adding at least one further ingredient, and iii.Mixing the powder and liquid obtained to form a uniform mixture.
 35. Themethod according to claim 34, wherein the further ingredient is selectedfrom fruits or vegetables.
 36. The method according to any of claims33-34, wherein the mixing is performed by shaking.
 37. A method forpreparing an instant beverage powder comprising BLG and at least oneoptional ingredient, said method comprising blending a dry BLG isolatewith the least one additional ingredient selected from the groupconsisting of vitamins, flavouring agent, colouring agent, minerals,sweeteners, antioxidants, food acid, lipids, carbohydrate, prebiotics,probiotics, anti-foaming agents and non-whey protein to obtain aninstant beverage powder.
 38. A method according to claim 37, wherein theBLG of the dry BLG isolate is coated with an organic acid selected fromthe group consisting of pyruvate, aconitate, citrate, iso-citrate,ketoglutarate, succinyl-CoA, succinate, fumarate, malate, oxaloacetate,tartrate, acetate, tannic acid, benzoic acid, maleic acid and lactate